supramolecular chemistry of beta– and gamma– … · supramolecular chemistry of ... important...
TRANSCRIPT
SUPRAMOLECULAR CHEMISTRY OF
BETA– AND GAMMA–
CYCLODEXTRIN DIMERS
Huy Tien Ngo
Thesis submitted for the degree of
Doctor of Philosophy
in
The University of Adelaide
School of Chemistry and Physics
October 2010
Huy Tien Ngo Chapter 4
135
CHAPTER 4
CONTROLLED SUPRAMOLECULAR
POLYMER ASSEMBLY OF 1-NAPHTHALENE
LABELLED POLY(ACRYLATE)S BY LINKED
- AND -CYCLODEXTRIN DIMERS
Huy Tien Ngo Chapter 4
136
4.1. Introduction
4.1.1. Controlled Supramolecular Assembly of Polymers and Hydrogels
Hydrogels are water-swollen polymeric networks containing cross-links of chemical or
physical nature. Research on new polymer hydrogels has become of increasing interest in
recent years because of potential applications in drug delivery, biosensing, tissue
engineering, functional nanodevices and biological coating technologies.1-6 In addition to
their biocompatibility characteristics, hydrogels can be tailored to be responsive to various
environmental stimuli during their development for different applications. It is therefore
important to understand and control the aqueous supramolecular assembly of polymers at
the molecular and macroscopic levels in order to produce hydrogels which exhibit
predictable and controllable character and which constitute new materials.
Figure 4.1 shows some possible interactions of substituted polymers in aqueous
solution. The substitution of hydrophobic entities (such as a 1-naphthyl group shown in A)
onto water soluble polymers solubilises these hydrophobes and facilitates studies of their
aggregation and complexation processes in aqueous solutions, which would normally be
otherwise inaccessible. This is likely to result in hydrophobe association and polymer
aggregation (B), enhancement of the viscosity of the polymer solution and possibly
formation of hydrogels. The addition of single water soluble hydrophobe receptors, such as
cyclodextrins (CDs) may lead to polymer disaggregation (C), which can be reversed in the
presence of competitive complexation by a second free hydrophobe species.7-9 On the other
hand, the addition of hydrophobe receptors substituted on water soluble polymers may
produce new entanglement and aggregation through hydrophobe–hydrophobe receptor
association (D) which leads to increased viscosity. This is exemplified by the 1:1 molar
ratio mixture of either CD or CD 3% randomly substituted poly(acrylate)s (PAA) with
octadecyl 3% substituted PAA,8,9 or with adamantyl 3% substituted PAA,10,11 which
resulted in up to a 100-fold increase in solution viscosity by comparison to that of the
individual polymers separately. Furthermore, the addition of dimeric hydrophobe receptors
is likely to lead to (dimeric hydrophobe receptor)(hydrophobe)2 complexation (E) which
can result in stronger polymer association and enhanced viscosity. For example, the
addition of a terephthalimide linked CD dimer to solutions of adamantyl-containing N,N’-
dimethylacrylamide or N-isopropylacrylamide copolymers increased the viscosity of the
solution dramatically to form stable gels within seconds.12 For these interactive systems
Huy Tien Ngo Chapter 4
137
control of the assembly process can be achieved in many ways such as either changing the
substituents or the extent of substitution, varying the host–guest molar ratio, adjusting the
tether length between polymer backbone and substituents, and changing polymer
concentration, ionic strength, pH or temperature.13-15
A B
C
D
E
hydrophobe
polymer backbone
hydrophobeassociation
hydrophobereceptor
hydrophone - single hydrophobe receptor association
hydrophone - single hydrophobe receptor association
hydrophone - bis hydrophobe receptor association
Figure 4.1. Various interactions of water soluble substituted polymers in water.
Huy Tien Ngo Chapter 4
138
4.1.2. Fluorescent Polymers
While the association of aliphatic hydrophobe substituted polymers is evident at the
macroscopic level through increased polymer viscosity, it is only through studies at the
molecular level that the detailed nature of this association may be understood in terms
of specific hydrophobe–hydrophobe or hydrophobe–receptor interactions. Thus, the
interactions between aromatic hydrophobe pairs in substituted polymers, which are
generally through intra- and inter-polymer – stacking, as exemplified by
anthracene,16 naphthalene17-24 or pyrene25-35 (Figure 4.2) are often evident through
changes in their UV–vis spectra and larger changes in their fluorescence spectra as a
consequence of exciton coupling. Furthermore, the hydrophobe–receptor interactions
may also lead to changes in the UV–vis or fluorescence spectra of the fluorophores,
which allow their quantification by spectrophotometric methods, such as steady–state
or time–resolved fluorescence spectroscopy.24,36-38
anthracene naphthalene pyrene
E-stilbene
NN
E-azobenzene
XNH2
RXNH2
XNH2 XNH2
RXNH2
Z-stilbene
N N
Z-azobenzeneR XNH2 R XNH2
Figure 4.2. Some aromatic hydrophobic amines suitable for substitution onto water soluble
polymers where R and X represent a variety of substituents.
Other hydrophobes such as azobenzene and stilbene (Figure 4.2) have also been
substituted onto polymers, and their light-responsive photoisomerisation used to actively
control the supramolecular assembly.39,40 It was shown that photoirradiation with UV or
visible light caused repetitive changes in the viscosity of a mixture of CD substituted
PAA (PAACD) and azobenzene substituted PAA (PAAC12Azo), consistent with the
azobenzene moiety photoisomerising from the Z- form to the E- form and vice versa.39
Huy Tien Ngo Chapter 4
139
4.1.3. Aims of This Study
The research described in this chapter aims to extend previous understanding in the
control of the supramolecular polymer assembly in water through the interactions of
hydrophobic modified poly(acrylate)s (PAAs) and -, - and -cyclodextrin either in their
free state or as substituents in PAA.7-11,13-15 The current study involves the preparation of
two new 3% randomly substituted 1-naphthyl-sulfonamide poly(acrylate)s with either a
diaminoethyl tether (PAA1NSen) or diaminohexyl tether (PAA1NShn) linking the 1-
naphthyl substituents and the polymer backbone (Figure 4.3).
The host–guest complexation between the 1-naphthyl-sulfonamide substituents of
PAA1NSen and PAA1NShn and the cyclodextrin hosts, CD and CD as well as their
succinamide–linked dimers, 33CD2suc, 66CD2suc, 33CD2suc and 66CD2suc (Figure
4.3) is investigated at the macroscopic level by rheology as well as at the molecular level
by 2D 1H NOESY NMR and fluorescence spectroscopy.
The experiments are expected to provide insight into the factors influencing
fluorescence properties, the impact of polymer substituent tether length, as well as the size
and geometry of the CD and CD dimers on the host–guest complexation behaviour and
the viscosity of the 1-naphthalene sbustituted poly(acrylate) solutions.
CO2- CO2
- CO2-
OHN
SO O
HN
O
OHHO
HO
O
n
1
234
56
=
CD, n = 7CD, n = 8
HN
O
O
NH
HN
O
O
NH
C6A C6A
C3A C3A
n = 7, 66CD2suc 33CD2sucn = 8, 66CD2suc 33CD2suc
m
PAA1NSen, m = 2PAA1NShn, m = 6(A)
(B)
(C)
Figure 4.3. Schematic structures of (A) PAA1NSen and PAA1NShn; (B) CD and CD;
and (C) succinamide–linked CD and CD dimers.
Huy Tien Ngo Chapter 4
140
4.2. 1-Naphthalene Randomly Substituted Poly(Acrylate)s
4.2.1. Synthesis
The preparation of the new 3% randomly substituted N-(2-aminoethyl)-1-naphthyl-
sulfonamide and N-(6-aminohexyl)-1-naphthyl-sulfonamide poly(acrylate)s, PAA1NSen
and PAA1NShn, required the preparation of 4-nitrophenyl naphthalene-1-sulfonate,
1NSNP, from 1-naphthalenesulfonyl chloride and 4-nitrophenol in dichloromethane by a
method similar to that reported in the literature.41 Treatment of 1NSNP with either 1,2-
diaminoethane in dichloromethane or 1,6-diaminohexane in N,N-dimethylformamide
afforded N-(2-aminoethyl)-1-naphthyl-sulfonamide, 1NSen, and N-(6-aminohexyl)-1-
naphthyl-sulfonamide, 1NShn, in moderate yields. A literature method7,8 was then adapted
to prepare the new 3% randomly substituted PAA1NSen and PAA1NShn in 80–90% yield
(Figure 4.4).
CO2- CO2
- CO2-
OHN
SO
OHN
SO O
Cl
S
O
O
NO2O
4-nitrophenol
H2N
S
O
O
NH
1,2-diaminoethane/DCMEt3N, DCM
or 1,6-diaminohexane/DMF1.
2.HC
H2C
COOH n
+2. NaOH
1. 60 oC/NMP, DCC 3%
H2N
S
O
O
NH
m
m
m
PAA1NSen, m = 2PAA1NShn, m = 6
1NSen, m = 21NShn, m = 6
1NSNP
Figure 4.4. Synthetic scheme for preparation of 3% randomly substituted PAA1NSen and
PAA1NShn.
The succinamide–linked CD and CD dimers, 33CD2suc, 66CD2suc, 33CD2suc
and 66CD2suc were synthesised as reported previously42 and in Chapter 3.43
Huy Tien Ngo Chapter 4
141
4.2.2. 1H NMR Spectra
The 1H NMR (300 MHz) spectra of 1NSNP, 1NSen and 1NShn in DMSO-d6 are shown
in Figure 4.5 and those of PAA, PAA1NSen and PAA1NShn in D2O are shown in Figure
4.6. The degree of substitution at the PAA carboxyl groups by 1-naphthyl-sulfonamide was
determined to be 3.0 ± 0.3 % from the 1H NMR spectra (Figure 4.6) according to the
method reported previously in the literature.7
Figure 4.5. 1H NMR (300 MHz) spectra of 1NSNP, 1NSen and 1NShn in DMSO-d6.
Huy Tien Ngo Chapter 4
142
CO2- CO2
- CO2-
OHN
SO O
HNm
PAA1NSen, m = 2PAA1NShn, m = 6
Figure 4.6. 1H NMR (300 Mz) spectra of PAA and 3% substituted PAA1NSen and
PAA1NShn in D2O.
Huy Tien Ngo Chapter 4
143
4.2.3. Photophysical Properties
Typical UV–vis absorbance and fluorescence spectra were recorded for ~0.013 wt%
~0.0033 wt% solutions of PAA1NSen and PAA1NShn, respectively and are shown in
Figure 4.7. The concentrations of both the 1NSen and 1NShn substituents were calculated
to be 4.0 × 10-5 and 1.0 × 10-5 mol dm-3 for the UV–vis and fluorescence spectra,
respectively, according to section 5.4.2.1. The variation of tether length linking the 1-
naphthyl substituents and the polymer backbone from two methylene groups in
PAA1NSen to six methylene groups in PAA1NShn appears to reduce the intensity of both
the UV–vis absorbance and emission of the 1-naphthyl groups. The fluorescence of
PAA1NShn appears to be quenched significantly as the tether length increases, consistent
with increased mobility of the 1-naphthyl groups as well as their increased exposure to the
bulk water molecules.
Figure 4.7. UV–vis absorbance spectra (left ordinate, blue lines) and relative fluorescence
spectra (right ordinate, red lines) of 3% substituted PAA1NSen (solid lines) and
PAA1NShn (dashed lines) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3). The
concentrations of the 1-naphthyl substituents are 4.0 × 10-5 and 1.0 × 10-5 mol dm-3 for
UV–vis and fluorescence, respectively.
0
100
200
300
400
500
0
1,000
2,000
3,000
4,000
5,000
6,000
250 300 350 400 450
Rel
ativ
e flu
ores
cenc
e (a
.u.)
Mol
ar a
bsor
ptio
n (m
ol-1
dm3
cm-1
)
Wavelength (nm)
Huy Tien Ngo Chapter 4
144
4.3. Rheological Determination of Viscosity
The host–guest interactions at the macroscopic level between the 1NSen and 1NShn
substituents on the poly(acrylate)s, PAA1NSen and PAA1NShn, and CD, CD and
their linked dimers, 33CD2suc, 33CD2suc, 66CD2suc and 66CD2suc, are expected
to affect the zero–shear viscosities of the solutions. Rheological measurements were
carried out on 5 wt% aqueous solutions of PAA1NSen and PAA1NShn alone and in the
presence of a 1:1 molar ratio of each of the CD hosts at pH 7.0 and [NaCl] = 0.10 mol
dm-3.
The variations of the viscosities of the five PAA1NSen and the five PAA1NShn
solutions with shear rate are shown in Figures 4.8 and 4.9. The variations of the
viscosities of PAA1NSen/CD, PAA1NSen/CD, PAA1NShn/CD and
PAA1NShn/CD solutions are very similar to those of PAA1NSen and PAA1NShn
solutions alone over the shear rate range studied, and thus are not shown. The zero–
shear viscosity variations, corresponding to the viscosities extrapolated from those
observed at the lowest shear rates, are shown graphically and numerically in Figure
4.10 and its caption.
Figure 4.8. Viscosity variations with shear rate of 5 wt% aqueous solutions of PAA1NSen
and PAA1NShn alone and in the presence of 66CD2suc and 66CD2suc at pH 7.0 and
[NaCl] = 0.10 mol dm-3 at 298.2 K. The concentrations of the CD and CD substituents in
the linked dimers are equal to those of the 1NSen or 1NShn poly(acrylate) substituents.
0.01
0.02
0.05
0.1
1 10 100 1000
PAA1NShn+66CD2suPAA1NShn+66CD2suPAA1NShnPAA1NSen+66CD2suPAA1NSen+66CD2suPAA1NSen
Shear Rate [1/s]
Vis
cosi
ty [P
as]
c c
c c
Huy Tien Ngo Chapter 4
145
Figure 4.9. Viscosity variations with shear rate of 5 wt% aqueous solutions of PAA1NSen and PAA1NShn alone and in the presence of 33CD2suc and 33CD2suc at pH 7.0 and [NaCl] = 0.10 mol dm-3 at 298.2 K. The concentrations of the CD and CD substituents in the linked dimers are equal to those of the 1NSen or 1NShn poly(acrylate) substituents.
Figure 4.10. Zero–shear viscosities of 5 wt% aqueous solutions of PAA1NSen (blue column) and PAA1NShn (red column) alone and in the presence of 33CD2suc, 33CD2suc, 66CD2suc and 66CD2suc at pH 7.0 and [NaCl] = 0.10 mol dm-3 at 298.2 K. The concentrations of the CD and CD substituents in the linked dimers are equal to those of the 1NSen or 1NShn poly(acrylate) substituents. A) PAA1NSen (0.0147) and PAA1NShn (0.0172); B) PAA1NSen/33CD2suc (0.0175) and PAA1NShn/33CD2suc (0.0206); C) PAA1NSen/33CD2suc (0.0159) and PAA1NShn/33CD2suc (0.0242); D) PAA1NSen/66CD2suc (0.0156) and PAA1NShn/66CD2suc (0.0315); E) PAA1NSen/66CD2suc (0.0164) and PAA1NShn/66CD2suc (0.0263), where the zero–shear viscosities (Pa·s) are shown in brackets.
0.01
0.02
0.05
0.1
1 10 100 1000
PAA1NShn+33CD2suPAA1NShn+33CD2suPAA1NShnPAA1NSen+33CD2suPAA1NSen+33CD2suPAA1NSen
Shear Rate [1/s]
Vis
cosi
ty [P
as]
0.000
0.005
0.010
0.015
0.020
0.025
0.030
0.035
A B C D E
Vis
cosi
ty [P
a·s]
c c
c c
Huy Tien Ngo Chapter 4
146
The viscosities of both PAA1NSen and PAA1NShn solutions alone (Figure 4.8) show
little variation with shear rate, consistent with very little or weak or both inter–polymer
strand cross–linking occurring in these solutions. The viscosity of PAA1NSen solution
(0.0147 Pa·s) is slightly smaller than that of PAA1NShn solution (0.0172 Pa·s), consistent
with the shorter ethyl tether of PAA1NSen restricting – association to a greater extent
than the longer hexyl tether of PAA1NShn. These values are similar to the viscosity of a 5
wt% solution of 3% substituted dodecyl poly(acrylate) (PAAC12), but about four orders of
magnitude smaller than that of a 5 wt% solution of 3% substituted octadecyl
poly(acrylate)s (PAAC18).13 This suggests that the intermolecular – interaction between
the 1-naphthyl substituents is of similar strength to hydrophobic association between the
dodecyl groups but much weaker than that between the octadecyl groups.
The viscosities of the PAA1NSen and PAA1NShn solutions in the presence of all four
linked CD and CD dimers (Figures 4.8 & 4.9) also show very little variation with shear
rate, consistent with little inter–strand cross–linking, similar to that occurring in
PAA1NSen and PAA1NShn solutions alone. The zero–shear viscosities of the PAA1NSen
solutions containing CD and CD dimers increase in the sequence of PAA1NSen
(0.0147) < PAA1NSen/66CD2suc (0.0156) < PAA1NSen/33CD2suc (0.0159) <
PAA1NSen/66CD2suc (0.0164) < PAA1NSen/33CD2suc (0.0175) (blue bars A, D, C, E,
B, Figure 4.10). Overall the increases in viscosity of PAA1NSen solutions containing CD
dimers over PAA1NSen solution alone are small, consistent with very little additional
cross–linking occurring through 1-naphthyl complexation by the CD dimers, which is
restricted by the short ethyl tether length. Most of the complexation is likely to occur
through single interaction between 1-naphthyl and a CD dimer (Figures 4.11a and 4.11b).
The zero–shear viscosities of the longer tether PAA1NShn solutions containing CD
and CD dimers increase in the sequence PAA1NShn (0.0172) < PAA1NShn/33CD2suc
(0.0206) < PAA1NShn/33CD2suc (0.0242) < PAA1NShn/66CD2suc (0.0263) <
PAA1NShn/66CD2suc (0.0315) (red bars A, B, C, E, D, Figure 4.10). The 1.2 to 1.8-fold
increase in viscosity of PAA1NShn/(CD dimer) solutions over PAA1NShn solution alone
are consistent with additional cross–linking caused by complexation of the 1NShn
substituents by the CD dimers (Figures 4.11c and 4.11d). This is consistent with the longer
hexyl tether allowing inter–strand complexation to form extra cross–links between polymer
strands. In addition, the larger increases in viscosity caused by 66CD2suc and 66CD2suc
Huy Tien Ngo Chapter 4
147
are consistent with the geometry of the 6,6–linked dimers favouring cross–linking over the
3,3–linked dimers. The highest viscosity of PAA1NShn/66CD2suc solution is consistent
with the wide rim of CD being the best fit for complexing the 1-naphthyl group.
CO2-
CO2-
CO2-
O
NH
SO2
NH
PAA1NSen orPAA1NShn
CO2-
CO2-
CO2-
O
NH
SO2
NH
-O2C
-O2C
-O2C
O
NH
O2S
HN
CO2-
CO2-
CO2-
O
NH
SO2
NH
-O2C
-O2C
-O2C
O
NH
O2S
HN
-O2C
-O2C
-O2C
O
NH
O2SHN CO2
-
CO2-
CO2-
O
NH
SO2
NH-O2C
-O2C
-O2C
O
NH
O2SHN
CO2-
CO2-
CO2-
O
NH
SO2
NH
-O2C
-O2C
-O2C
O
NH
O2SHN
CO2-
CO2-
CO2-
O
NH
SO2
NH
-O2C
-O2C
-O2C
O
NH
O2SHN
HN
O
HN
O
HN
O
HN
O
+ 66CD2suc or+ 66CD2suc
+ 33CD2suc or+ 33CD2suc
HN
O
HN
O
HN
O
HN
O
HN
O
HN
O
HN
O
HN
O
a) PAA1NSen/66CD2suc or PAA1NSen/66CD2suc
b) PAA1NSen/33CD2suc or PAA1NSen/33CD2suc
c) PAA1NShn/66CD2suc or PAA1NShn/66CD2suc
d) PAA1NShn/33CD2suc or PAA1NShn/33CD2suc
Figure 4.11. Representations of the single complexation of PAA1NSen by a) 66CD2suc
and 66CD2suc and b) 33CD2suc and 33CD2suc and the single and double complexation
of PAA1NShn by c) 66CD2suc and 66CD2suc and d) 33CD2suc and 33CD2suc.
Huy Tien Ngo Chapter 4
148
4.4. 2D 1H NOESY NMR Studies
The host–guest interactions between the 1-naphthyl substituents of PAA1NSen and
PAA1NShn and CD, CD and their succinamide-linked dimers, 33CD2suc,
66CD2suc, 33CD2suc and 66CD2suc, were studied at the molecular level through
2D 1H NOESY NMR spectroscopy. The 2D 1H NOESY NMR (600 MHz, 300 ms
mixing time) spectra were recorded on 1.43 wt% PAA1NSen and PAA1NShn solutions
(10 mg in 0.70 cm3) in D2O at pD = 7.0 with 0.10 mol dm-3 NaCl at 298.2 K. The
corresponding concentrations of the 1NSen or 1NShn substituents were 3.00 × 10-3 mol
dm-3 and 2.94 × 10-3 mol dm-3, respectively. In each system, the same concentration of
CD, CD or a linked CD dimer to that of either the 1NSen or 1NShn substituents was
present in the solution.
The 2D 1H NOESY NMR spectra for the PAA1NSen solutions with CD, CD and
the linked CD dimer hosts are shown in Figures 4.12–4.17 in section 4.4.1 and the
analogous spectra for the PAA1NShn solutions are shown in Figures 4.18–4.23 in
section 4.4.2. In each system, strong cross–peaks arising from dipolar interactions
between the aromatic protons of the 1-naphthyl groups and the H3,5,6 annular protons of
either CD or CD or their linked dimers are observed. In addition, there are cross–
peaks arising from interactions between the protons of the hexyl tether in PAA1NShn
and the H3,5,6 annular protons of either CD or CD or their linked CD dimers. On the
other hand, cross–peaks arising from interactions of the ethyl tether protons in
PAA1NSen are not observed. These data are consistent with both the hexyl tether and
the 1-naphthyl group of PAA1NShn complexing within CD or CD or their linked
dimers, whereas only the 1-naphthyl group of PAA1NSen is complexed within the CD
hosts. The relative strength of cross–peaks between PAA1NSen aromatic protons and
H3,5,6 of CD and CD dimers increases in the sequence 33CD2suc < 66CD2suc <
33CD2suc < 66CD2suc; and for PAA1NShn the sequence is 33CD2suc < 33CD2suc
< 66CD2suc < 66CD2suc for aromatic 1-naphthyl protons and 33CD2suc <
66CD2suc < 33CD2suc < 66CD2suc for the hexyl tether protons.
Although the 1H NMR data are consistent with host–guest complexation occurring,
they do not distinguish between single substituent complexation and the simultaneous
complexation of 1NSen and 1NShn substituents by the CD or CD dimers to form
Huy Tien Ngo Chapter 4
149
inter–polymer strand cross–links. Nevertheless, both 1H NMR and viscosity data
suggest that the ability to form additional inter–strand cross–links by host–guest
complexation depends upon the tether length of the substituted polymers, as well as the
size and geometry of the CD and CD dimers.
4.4.1. PAA1NSen
Figure 4.12. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar CD in D2O at pD 7.0 with 0.10
mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed in the
rectangle arise from interaction between the annular CD protons H3,5,6 and the naphthyl
protons of the 1NSen substituent. Above: model representation of the complexation
between CD and the 1NSen substituent (blue).
Huy Tien Ngo Chapter 4
150
Figure 4.13. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar CD in D2O at pD 7.0 with 0.10
mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed in the
rectangle arise from interaction between the annular CD protons H3,5,6 and the naphthyl
protons of the 1NSen substituent. Above: model representation of the complexation
between CD and the 1NSen substituent (blue).
Huy Tien Ngo Chapter 4
151
Figure 4.14. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar 33CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangle arise from interaction between the annular CD protons H3,5,6 and the
naphthyl protons of the 1NSen substituent. Above: model representation of the
complexation between 33CD2suc and the 1NSen substituent (blue).
Huy Tien Ngo Chapter 4
152
Figure 4.15. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar 66CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangle arise from interaction between the annular CD protons H3,5,6 and the
naphthyl protons of the 1NSen substituent. Above: model representation of the
complexation between 66CD2suc and the 1NSen substituent (blue).
Huy Tien Ngo Chapter 4
153
Figure 4.16. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar 33CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangle arise from interaction between the annular CD protons H3,5,6 and the
naphthyl protons of the 1NSen substituent. Above: model representation of the
complexation between 33CD2suc and the 1NSen substituent (blue).
Huy Tien Ngo Chapter 4
154
Figure 4.17. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NSen
(1.43 wt%, [1NSen] = 3.0 x 10-3 mol dm-3) and equimolar 66CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangle arise from interaction between the annular CD protons H3,5,6 and the
naphthyl protons of the 1NSen substituent. Above: model representation of the
complexation between 66CD2suc and the 1NSen substituent (blue).
CD H2-6
CD H1
Huy Tien Ngo Chapter 4
155
4.4.2. PAA1NShn
Figure 4.18. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar CD in D2O at pD 7.0 with
0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed in the
rectangles arise from interaction between the annular CD protons H3,5,6 and the naphthyl
and hn CH2 protons of the 1NShn substituent. Above: model representation of the
complexation between CD and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
156
Figure 4.19. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar CD in D2O at pD 7.0 with
0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed in the
rectangles arise from interaction between the annular CD protons H3,5,6 and the naphthyl
and hn CH2 protons of the 1NShn substituent. Above: model representation of the
complexation between CD and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
157
Figure 4.20. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar 33CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangles arise from interaction between the annular CD protons H3,5,6 and the
naphthyl, hn N-CH2 and hn CH2 protons of the 1NShn substituent. Above: model
representation of the complexation between 33CD2suc and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
158
Figure 4.21. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar 66CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangles arise from interaction between the annular CD protons H3,5,6 and the
naphthyl, hn N-CH2 and hn CH2 protons of the 1NShn substituent. Above: model
representation of the complexation between 66CD2suc and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
159
Figure 4.22. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar 33CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangles arise from interaction between the annular CD protons H3,5,6 and the
naphthyl, hn N-CH2 and hn CH2 protons of the 1NShn substituent. Above: model
representation of the complexation between 33CD2suc and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
160
Figure 4.23. 2D 1H NOESY NMR (600 MHz) spectrum of 3% substituted PAA1NShn
(1.43 wt%, [1NShn] = 2.94 x 10-3 mol dm-3) and equimolar 66CD2suc in D2O at pD 7.0
with 0.10 mol dm-3 NaCl at 298.2 K with a mixing time of 300 ms. Cross–peaks enclosed
in the rectangles arise from interaction between the annular CD protons H3,5,6 and the
naphthyl and hn CH2 protons of the 1NShn substituent. Above: model representation of the
complexation between 66CD2suc and the 1NShn substituent (blue).
Huy Tien Ngo Chapter 4
161
4.5. Fluorimetric Determination of Host–Guest Complexation
The host–guest interactions between the 1-naphthyl substituents of PAA1NSen and
PAA1NShn and CD, CD and their succinamide-linked dimers, 33CD2suc,
66CD2suc, 33CD2suc and 66CD2suc, were further studied at the molecular level by
fluorescence spectroscopy. Fluorescence spectra were recorded for 0.0033 wt%
PAA1NSen and 0.0034 wt% PAA1NShn solutions, respectively (the calculated
concentrations of both the 1NSen and 1NShn substituents are 1.0 × 10-5 mol dm-3) in
pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K. At these low concentrations,
the optical density of the 1-naphthyl substituents is below 0.05 and therefore any inner–
filter effects on fluorescence are negligible.44
Variations in the fluorescence spectra were monitored as the samples of either 0.0033
wt% PAA1NSen or 0.0034 wt% PAA1NShn solutions were sequentially diluted with
0.050 cm3 aliquots of CD solution (1.06 × 10-2 mol dm-3), CD solution (4.96 × 10-2 mol
dm-3), 33CD2suc solution (2.49 × 10-3 mol dm-3), 66CD2suc solution (2.31 × 10-3 mol
dm-3), 33CD2suc solution (2.63 × 10-3 mol dm-3) or 66CD2suc solution (2.49 × 10-3 mol
dm-3) over the range 300–550 nm in 0.5 nm intervals. The fluorescence spectra and fittings
for PAA1NSen with CD, CD and their dimers are shown in section 4.5.1 and analogous
spectra and fittings for PAA1NShn are shown in section 4.5.2.
At the low concentrations of the fluorescence studies, aggregation of 1-naphthyl groups
appears to be negligible, since no deviation from Beer’s law in the our absorption spectra
of either the PAA1NSen or PAA1NShn was observed as the concentration of the 1-
naphthyl substituents was diluted from 1.0 × 10-4 mol dm-3 to 4.0 × 10-5 mol dm-3.
Therefore, the most likely equilibria in the solutions will be the formation of either 1:1 or
1:2 host–guest complex or both between the 1-naphthyl substituents and CD or CD or a
CD or CD dimer, as exemplified by Eqns. 4.1 and 4.2 for the complexation between
66CD2suc and 1NShn. Analogous equations apply for the other PAA1NSen and
PAA1NShn systems.
66CD2suc + 1NShn 66CD2suc.1NShn (4.1)K1
66CD2suc.1NShn + 1NShn 66CD2suc.(1NShn)2 (4.2)K2
Huy Tien Ngo Chapter 4
162
The observed fluorescence intensity, IF, at any given wavelength is equal to the mole
fraction–weighted sum of the fluorescence intensities of the free and 1:1 and 1:2
complexed 1NShn species, as shown in Eqn. 4.3.
IF = IF(1NShn)([1NShn]/[1NShn]total) + IFCD2suc.1NShn)([66CD2suc.1NShn]/[1NShn]total)
+ IFCD2suc.(1NShn)2)([66CD2suc.(1NShn)2]/[1NShn]total) (4.3)
The stepwise stability constants, K1 and K2, for the 1:1 and 1:2 host–guest complexes,
respectively, are defined by the following equations:
K1 = [66CD2suc.1NShn]/([66CD2suc][1NShn]) (4.4)
K2 = [66CD2suc.(1NShn)2]/([66CD2suc.1NShn][1NShn]) (4.5)
Except for the systems of PAA1NSen/66CD2suc (Figure 4.29), PAA1NShn/CD
(Figure 4.30), PAA1NShn/33CD2suc (Figure 4.32), PAA1NShn/66CD2suc (Figure
4.33) and PAA1NShn/66CD2suc (Figure 4.35), there is a decrease in both the observed
fluorescence intensity and the relative fluorescence intensity, which is the fluorescence
intensity corrected to the same concentration of the 1-naphthyl substituents relative to the
fluorescence intensity of the 1-naphthyl substituents alone (1.0 × 10-5 mol dm-3) as the ratio
of [CD host]total/[1NS guest]total increases. A slight blue-shift of emission maxima occurs in
all systems. An algorithm for the formation of 1:1 host–guest complex analogous to Eqn.
4.3, with the absence of the third right-hand term, best–fits the data in ranges of
wavelengths, where significant fluorescence changes occur, as indicated in the figure
captions, to yield the complexation constants, K1, which appear in Table 4.1. In most cases,
good fits of the fluorescence data were obtained, except for the PAA1Nsen/66CD2suc
system, where the fluorescence variation was relatively small resulting in a derived K1
value with ~30 % error.
In the cases of PAA1NSen/66CD2suc (Figure 4.29), PAA1NShn/66CD2suc (Figure
4.33) and PAA1NShn/66CD2suc (Figure 4.35) systems, an increase in the relative
fluorescence intensity was observed coupled with a much stronger blue-shift of emission
maxima as each CD dimer host was added. For these complexes, the algorithm for the
formation of 1:1 host–guest complex did not fit the fluorescence data, instead an algorithm
analogous to Eqn. 4.3 for the formation of both 1:1 and 1:2 host–guest complexes best–fits
the data to yield the stepwise complexation constants, K1 and K2, which also appear in
Huy Tien Ngo Chapter 4
163
Table 4.1. Overall, the errors in the fittings of K1 and K2 for these systems are large (10 –
25 %) over the ranges of the linked CD dimer concentrations being studied.
In the case of PAA1NShn/33CD2suc system (Figure 4.32), the observed fluorescence
decreased but the relative fluorescence remained little changed upon serially addition of
33CD2suc solution. Consequently, a reliable fitting to the experimental data could not be
obtained to derive the complexation constant, K1, consistent with little complexation
occurring.
Huy Tien Ngo Chapter 4
164
4.5.1. Fluorimetric Titrations of PAA1NSen Complexation
Figure 4.24. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of CD solution (1.06 × 10-2 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [CD]total/[1NSen]total increases. max= 360 nm and 359 nm for the free and complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–420 nm for both cases. Bottom: Speciation relative to [1NSen]total, curve a is the percentage of free [1NSen] and curve b is the percentage of [CD.1NSen].
0
100
200
300
400
500
300 350 400 450 500
Fluo
resc
ence
inte
nsity
(a.u
.)
Wavelength (nm)
0.90
0.95
1.00
220
320
420
0 200 400 600 800
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[CD]total/[1NSen]total
0
20
40
60
80
100
0 200 400 600 800
% s
peci
atio
n
[CD]total/[1NSen]total
a
b
Huy Tien Ngo Chapter 4
165
Figure 4.25. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of CD solution (4.96 × 10-2 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [CD]total/[1NSen]total increases. max= 360 nm and 355 nm for the free and complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–420 nm for both cases. Bottom: Speciation relative to [1NSen]total, curve a is the percentage of free [1NSen] and curve b is the percentage of [CD.1NSen].
0
100
200
300
400
500
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.70
0.80
0.90
1.00
180
280
380
0 1000 2000 3000
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[CD]total/[1NSen]total
0
20
40
60
80
100
0 1000 2000 3000
% s
peci
atio
n
[CD]total/[1NSen]total
a
b
Huy Tien Ngo Chapter 4
166
Figure 4.26. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 33CD2suc solution (2.49 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [33CD2suc]total/[1NSen]total increases. max= 360 nm and 356 nm for the free and complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–410 nm for both cases. Bottom: Speciation relative to [1NSen]total, curve a is the percentage of free [1NSen] and curve b is the percentage of [33CD2suc.1NSen].
0
100
200
300
400
500
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.85
0.90
0.95
1.00
200
300
400
0 50 100 150
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[33CD2suc]total/[1NSen]total
0
20
40
60
80
100
0 50 100 150
% s
peci
atio
n
[33CD2suc]total/[1NSen]total
a
b
Huy Tien Ngo Chapter 4
167
Figure 4.27. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 66CD2suc solution (2.31 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [66CD2suc]total/[1NSen]total increases. max= 360 nm and 354 nm for the free and complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the tentative best fit of the algorithm for a 1:1 complexation model in the range 330–410 nm for both cases. Bottom: speciation relative to [1NSen]total, curve a is the percentage of free [1NSen] and curve b is the percentage of [66CD2suc.1NSen].
0
100
200
300
400
500
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.96
0.98
1
1.02
200
300
400
500
0 25 50 75 100 125 150
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[66CD2suc]total/[1NSen]total
0
20
40
60
80
100
0 25 50 75 100 125 150
% s
peci
atio
n
[66CD2suc]total/[1NSen]total
a
b
Huy Tien Ngo Chapter 4
168
Figure 4.28. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 33CD2suc solution (2.63 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [33CD2suc]total/[1NSen]total increases. max= 360 nm and 359 nm for the free and complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–410 nm for both cases. Bottom: speciation relative to [1NSen]total, curve a is the percentage of free [1NSen] and curve b is the percentage of [33CD2suc.1NSen].
0
100
200
300
400
500
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.00
0.50
1.00
0
100
200
300
400
0 50 100 150
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[33CD2suc]total/[1NSen]total
0
20
40
60
80
100
0 50 100 150
% s
peci
atio
n
[33CD2suc]total/[1NSen]total
a
b
Huy Tien Ngo Chapter 4
169
Figure 4.29. Top: Variation in the emission spectra of 0.0033 wt% PAA1NSen solution ([1NSen] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 66CD2suc solution (2.52 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [66CD2suc]total/[1NSen]total increases. max= 360 nm, 347 nm and 349 nm for the free, 1:1 and 1:2 host–guest complexed 1NSen, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 345 nm. The solid curves represent the best fit of the algorithm incorporating 1:1 and 1:2 host–guest complexation models in the range 320–480 nm for both cases. Bottom: speciation relative to [1NSen]total, curve a is the percentage of free [1NSen], curve b is the percentage of [66CD2suc.1NSen] and curve c is twice the percentage of [66CD2suc.(1NSen)2].
0
100
200
300
400
500
600
700
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
1.00
2.00
3.00
370
470
570
670
0 50 100 150
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[66CD2suc]total/[1NSen]total
0
20
40
60
80
100
0 50 100 150
% s
peci
atio
n
[66CD2suc]total/[1NSen]total
a
bc
Huy Tien Ngo Chapter 4
170
4.5.2. Fluorimetric Titrations of PAA1NShn Complexation
Figure 4.30. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution ([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.350 cm3 initially then 0.050 cm3 each) of CD solution (1.06 × 10-2 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [CD]total/[1NShn]total increases. max= 360 nm and 350 nm for the free and complexed 1NShn, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 355 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–430 nm for both cases. Bottom: Speciation relative to [1NShn]total, curve a is the percentage of free [1NShn] and curve b is the percentage of [CD.1NShn].
0
50
100
150
200
310 360 410 460
Fluo
resc
ence
inte
nsity
(a.u
.)
Wavelength (nm)
1.00
1.05
1.10
120
140
160
185 385 585 785
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[CD]total/[1NShn]total
0
20
40
60
80
100
185 385 585 785
% s
peci
atio
n
[CD]total/[1NShn]total
a
b
Huy Tien Ngo Chapter 4
171
Figure 4.31. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution ([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of CD solution (4.96 × 10-2 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [CD]total/[1NShn]total increases. max= 360 nm and 346 nm for the free and complexed 1NShn, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 355 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 330–430 nm for both cases. Bottom: Speciation relative to [1NShn]total, curve a is the percentage of free [1NShn] and curve b is the percentage of [CD.1NShn].
0
50
100
150
200
300 350 400 450 500Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.70
0.80
0.90
1.00
80
100
120
140
160
180
0 1000 2000 3000
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[CD]total/[1NShn]total
0
20
40
60
80
100
0 1000 2000 3000
% s
peci
atio
n
[CD]total/[1NShn]total
a
b
Huy Tien Ngo Chapter 4
172
Figure 4.32. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution
([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K
upon sequential injection (0.050 cm3 each) of 33CD2suc solution (2.13 × 10-3 mol dm-3).
Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The
arrows indicate the direction of fluorescence changes as the ratio of
[33CD2suc]total/[1NShn]total increases. Bottom: Variation in the observed fluorescence
(left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 355 nm.
Reliable fittings of either 1:1 or 1:2 host–guest complexation model or both to the
experimental data could not be obtained.
0
50
100
150
200
310 360 410 460 510
Fluo
resc
ence
inte
nsity
(a.u
.)
Wavelength (nm)
0.90
1.00
1.10
100
120
140
160
180
0 25 50 75 100 125
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[33CD2suc]total/[1NShn]total
Huy Tien Ngo Chapter 4
173
Figure 4.33. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution ([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 66CD2suc solution (2.31 × 10-3 mol dm-3). Excitation wavelength ex = 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [66CD2suc]total/[1NShn]total increases. max = 360 nm, 351 nm and 354 nm for the free, 1:1 and 1:2 complexed 1NShn, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 355 nm. The solid curves represent the best fit of the algorithm incorporating 1:1 and 1:2 host–guest complexation models in the range 320–400 nm for both cases. Bottom: speciation relative to [1NShn]total, curve a is the percentage of free [1NShn], curve b is the percentage of [66CD2suc.1NShn] and curve c is twice the percentage of [66CD2suc.(1NShn)2].
0
50
100
150
200
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
1.00
1.05
1.10
120
140
160
180
0 25 50 75 100 125
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[66CD2suc]total/[1NShn]total
0
20
40
60
80
100
0 25 50 75 100 125
% s
peci
atio
n
[66CD2suc]total/[1NShn]total
a
bc
Huy Tien Ngo Chapter 4
174
Figure 4.34. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution ([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 33CD2suc solution (2.63 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [33CD2suc]total/[1NShn]total increases. max= 360 nm and 354 nm for the free and complexed 1NShn, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 360 nm. The solid curves represent the best fit of the algorithm for a 1:1 complexation model in the range 320–410 nm for both cases. Bottom: Speciation relative to [1NShn]total, curve a is the percentage of free [1NShn] and curve b is the percentage of [33CD2suc.1NShn].
0
50
100
150
200
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
0.20
0.60
1.00
20
70
120
170
0 50 100
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[33CD2suc]total/[1NShn]total
0
30
60
90
0 30 60 90 120
Fluo
resc
ence
inte
nsity
(a.u
.)
[33CD2suc]total/[1NShn]total
a
b
Huy Tien Ngo Chapter 4
175
Figure 4.35. Top: Variation in the emission spectra of 0.0034 wt% PAA1NShn solution ([1NShn] = 1.0 × 10-5 mol dm-3) in pH 7.0 phosphate buffer (I = 0.10 mol dm-3) at 298.2 K upon sequential injection (0.050 cm3 each) of 66CD2suc solution (2.52 × 10-3 mol dm-3). Excitation wavelength ex= 290 nm with both excitation and emission slits of 5 nm. The arrows indicate the direction of fluorescence changes as the ratio of [66CD2suc]total/[1NShn]total increases. max= 360 nm, 342 nm and 351 nm for the free, 1:1 and 1:2 complexed 1NShn, respectively. Middle: Variation in the observed fluorescence (left ordinate, circles) and relative fluorescence (right ordinate, triangles) at 355 nm. The solid curve a represents the best fit of the algorithm incorporating 1:1 and 1:2 host–guest complexation models in the range 320–430 nm for both cases. Bottom: speciation relative to [1NShn]total, curve a is the percentage of free [1NShn], curve b is the percentage of [66CD2suc.1NShn] and curve c is twice the percentage of [66CD2suc.(1NShn)2].
0
200
400
600
310 360 410 460 510Fl
uore
scen
ce in
tens
ity (a
.u.)
Wavelength (nm)
1.00
2.00
3.00
4.00
5.00
6.00
170
270
370
470
570
0 50 100 150
Rel
ativ
e flu
ores
cenc
e
Fluo
resc
ence
inte
nsity
(a.u
.)
[66CD2suc]total/[1NShn]total
0
20
40
60
80
100
0 50 100 150
% s
peci
atio
n
[66CD2suc]total/[1NShn]total
a
bc
Huy Tien Ngo Chapter 4
176
Table 4.1 summarises the stepwise stability constants, K1 and K2, for the 1:1 and 1:2
host–guest complexes of the 1NSen and 1NShn poly(acrylate) substituents by CD, CD
and their succinamide–linked dimers at the molecular level by fluorescence spectroscopy.
The results show differing fluorescence variations of PAA1NSen and PAA1NShn upon
complexation with CD, CD and the CD dimers, consistent with the variation in tether
length between ethyl and hexyl, as well as in size and geometry of CD, CD and the CD
dimer hosts.
Table 4.1. Stepwise equilibrium constants, K1 and K2, for the 1:1 and 1:2 host–guest
complexation of 3% randomly substituted PAA1NShn and PAA1NSen by CD, CD and
their succinamide–linked dimers, determined by fluorimetric titrations in pH 7.0 phosphate
buffer (I = 0.10 mol dm-3) at 298.2 K.
Host
PAA1NSen PAA1NShn
K1
dm3 mol-1
10-4 × K2
dm3 mol-1
K1
dm3 mol-1
10-4 × K2
dm3 mol-1
CD 440 ± 20 – 60 ± 10 –
33CD2suc 420 ± 20 – very small –
66CD2suc 120 ± 40 – 160 ± 40 7.9 ± 0.8
CD ~20 – 330 ± 15 –
33CD2suc 1150 ± 60 – 1230 ± 60 –
66CD2suc ~20 30 ± 6 ~20 37 ± 7
The fluorescence of PAA1NSen and PAA1NShn show contrasting behaviour upon their
complexation by the native CD and CD. While CD complexes PAA1NSen much more
strongly than CD does (~22–fold) as evident by larger fluorescence variation, the longer
tethered and weakly fluorescent PAA1NShn is complexed 5.5–fold more strongly by CD
than it is by CD. This is consistent with the 1-naphthyl moiety of the short tethered
PAA1NSen fitting better to the CD annulus than the larger CD annulus. On the other
hand, the longer hexyl tether of PAA1NShn acts as a “space competitor” to compete with
1-naphthyl moiety in complexing within the CD annulus, but acts as “space regulator” to
optimise the fit of 1-naphthyl to the bigger CD annulus. The competition between the
Huy Tien Ngo Chapter 4
177
hexyl and 1-naphthyl moieties in complexing within CD is consistent with the much
smaller fluorescence variation of PAA1NShn induced by CD complexation (Figure 4.30)
by comparison with that of PAA1NSen (Figure 4.24).
In addition to the effects of polymer tether lengths and CD and CD annular sizes,
differing geometries between the 3,3–linked and the 6,6–linked CD and CD dimers
provide a third controlling factor over host–guest complexation. Both 33CD2suc and
33CD2suc complex PAA1NSen and PAA1NShn in a 1:1 stoichiometry, in a similar way
to that which native CD and CD do. With the narrow end of two smaller CD annuli
pointing outward, 33CD2suc only complexes PAA1NSen moderately, while the
competition of the hexyl tether in PAA1NShn leads to very weak interaction of the 1-
naphthyl moiety with 33CD2suc (Figure 4.32). On the other hand, the larger CD annuli
of 33CD2suc allow stronger complexation to both PAA1NSen and PAA1NShn (Figures
4.28 and 4.34). Neither 33CD2suc nor 33CD2suc show inter–polymer strand cross–
linking through 1:2 host–guest complexation with either PAA1NSen or PAA1NShn. This
is consistent with two CD or CD annuli being of relatively close proximity to each other
due to the inversion of the C2A and C3
A carbons on each of the CD or CD altropyranose
units in these dimers, thus restricting intermolecular cross–linking.
In contrast, 66CD2suc and 66CD2suc show additional 1:2 host–guest complexation
with both PAA1NSen and PAA1NShn, except for the 66CD2suc/PAA1NSen system
where only the 1:1 host–guest complex is observed. For these systems, the stepwise
stability constants, K2, of the 1:2 complexes are about 2.7 to ~4.2 orders of magnitude
higher than K1 of the 1:1 complexes (Table 4.1), coupled with opposite fluorescence
behaviour as compared to the other systems (fluorescence enhancing instead of quenching,
Figures 4.29 & 4.35). This opposite fluorescence behaviour can be attributed to additional
- interaction of two adjacent 1-naphthyl groups induced by complexation by either
66CD2suc or 66CD2suc. This phenomenon has been observed previously, where induced
dimerisation of CD-appended anthracene occurs within the CD annulus, with the
stepwise 1:2 host–guest complexation constant, K2, of almost two orders of magnitude
higher than the 1:1 complexation constant, K1.45 The 1:2 host–guest complexation of
PAA1NShn by 66CD2suc and 66CD2suc appears to provide additional inter–polymer
strand cross-links which lead to enhanced viscosities of these solutions as shown in section
Huy Tien Ngo Chapter 4
178
4.3 (red bars D and E, Figure 4.10, page 145). On the other hand, 1:2 host–guest
complexation between 66CD2suc and PAA1NSen do not greatly enhance solution
viscosity, consistent with the short ethyl tether restricting inter–strand cross–linking (blue
bar E, Figure 4.10, page 145). Overall, the fluorescence data agree with the 1H NMR and
viscosity data in that the ability to form additional inter–strand cross–links through host–
guest complexation depends upon both tether length of the substituted polymers and the
size and geometry of the CD and CD dimers.
4.6. Conclusion
Two new 3% randomly substituted poly(acrylate)s labelled with 1-naphthalene side
groups through either a two methylene tether, PAA1NSen, or six methylene tether,
PAA1NShn, have been synthesised and studied for their polymer network assembly in
aqueous solution. The zero–shear viscosities of 5 wt% solutions of both PAA1NSen and
PAA1NShn are similar to each other and similar to that of 5 wt% solution of 3% randomly
substituted dodecyl poly(acrylate) (PAAC12), but are several orders of magnitude smaller
than that of 5 wt% solution of 3% randomly substituted octadecyl poly(acrylate)
(PAAC18). This is consistent with - stacking between inter–polymer 1-naphthyl groups,
being the main force for the assembly, having similar strength to the hydrophobic
interaction between dodecyl substituents but much weaker than hydrophobic interaction
between the longer octadecyl substituents.
The patterns of the relative magnitudes of the effects of four CD and CD dimers,
33CD2suc, 33CD2suc, 66CD2suc and 66CD2suc, on the polymer network assembly of
PAA1NSen and PAA1NShn have been studied at the macroscopic level by viscosity and at
the molecular level by 1H NOESY NMR and fluorescence spectroscopy. The three sets of
data are together consistent with a combination of the tether lengths linking the 1-naphthyl
substituents to the polymer backbone, the sizes of the CD and CD annuli and the
geometries of the linked CD and CD dimers controlling the extent and strength of the
inter–polymer strand cross–links formed through 1-naphthyl substituent aggregation and
host–guest complexation. This has provided insight for the design of new aqueous polymer
networks and hydrogels with potential for practical application.
Huy Tien Ngo Chapter 4
179
4.7. References
1. Peppas, N. A.; Huang, Y.; Torres-Lugo, M.; Ward, J. H.; Zhang, J., Annu. Rev.
Biomed. Eng. 2000, 2, 9-29.
2. Lee, K. Y.; Mooney, D. J., Chem. Rev. 2001, 101, 1869-1879.
3. Nowak, A. P.; Breedveld, V.; Pakstis, L.; Ozbas, B.; Pine, D. J.; Pochan, D.;
Deming, T. J., Nature 2002, 417, 424-428.
4. Sangeetha, N. M.; Maitra, U., Chem. Soc. Rev. 2005, 34, 821-836.
5. Hendrickson, G. R.; Lyon, L. A., Soft Matter 2009, 5, 29-35.
6. Huebsch, N.; Mooney, D. J., Nature 2009, 462, 426-432.
7. Guo, X.; Abdala, A. A.; May, B. L.; Lincoln, S. F.; Khan, S. A.; Prud'homme, R. K.,
Macromolecules 2005, 38, 3037-3040.
8. Guo, X.; Abdala, A. A.; May, B. L.; Lincoln, S. F.; Khan, S. A.; Prud'homme, R. K.,
Polymer 2006, 47, 2976-2983.
9. Li, L.; Guo, X.; Fu, L.; Prud'homme, R. K.; Lincoln, S. F., Langmuir 2008, 24, 8290-
8296.
10. Guo, X.; Wang, J.; Li, L.; Pacheco, C. R.; Fu, L.; Prud'homme, R. K.; Lincoln, S. F.,
PMSE Preprints 2007, 97, 543-544.
11. Li, L.; Guo, X.; Wang, J.; Liu, P.; Prud'homme, R. K.; May, B. L.; Lincoln, S. F.,
Macromolecules 2008, 41, 8677-8681.
12. Kretschmann, O.; Choi, S. W.; Miyauchi, M.; Tomatsu, I.; Harada, A.; Ritter, H.,
Angew. Chem. Int. Ed. 2006, 45, 4361-4365.
13. Wang, J.; Li, L.; Ke, H.; Liu, P.; Zheng, L.; Guo, X.; Lincoln, S. F., Asia-Pac. J.
Chem. Eng. 2009, 4, 537-543.
14. Wang, J.; Pham, D.-T.; Guo, X.; Li, L.; Lincoln, S. F.; Luo, Z.; Ke, H.; Zheng, L.;
Prud'homme, R. K., Ind. Eng. Chem. Res. 2010, 49, 609-612.
Huy Tien Ngo Chapter 4
180
15. Guo, X.; Wang, J.; Li, L.; Pham, D.-T.; Clements, P.; Lincoln, S. F.; May, B. L.;
Chen, Q.; Zheng, L.; Prud'homme, R. K., Macromol. Rapid Commun. 2010, 31, 300-
304.
16. Clements, J. H.; Webber, S. E., Macromolecules 2004, 37, 1531-1536.
17. Cao, T.; Yin, W.; Webber, S. E., Macromolecules 1994, 27, 7459-7464.
18. Premachandran, R. S.; Banerjee, S.; Wu, X.-K.; John, V. T.; McPherson, G. L.,
Macromolecules 1996, 29, 6452-6460.
19. Hu, Y.; Smith, G. L.; Richardson, M. F.; McCormick, C. L., Macromolecules 1997,
30, 3526-3537.
20. Anghel, D. F.; Alderson, V.; Winnik, F. M.; Masanobu, M.; Morishima, Y., Polymer
1998, 39, 3035-3044.
21. Costa, T.; Miguel, M. G.; Lindman, B.; Schillén, K.; Lima, J. C.; Seixas de Melo, J.,
J. Phys. Chem. B 2005, 109, 3243-3251.
22. Costa, T.; Miguel, M. G.; Lindman, B.; Schillén, K.; Seixas de Melo, J., J. Phys.
Chem. B 2005, 109, 11478-11492.
23. Schillén, K.; Anghel, D. F.; Miguel, M. G.; Lindman, B., Langmuir 2000, 16, 10528-
10539.
24. Harada, A.; Ito, F.; Tomatsu, I.; Shimoda, K.; Hashidzume, A.; Takashima, Y.;
Yamaguchi, H.; Kamitori, S., J. Photochem. Photobiol. A: Chem. 2006, 179, 13-19.
25. Prazeres, T. J. V.; Beingessner, R.; Duhamel, J., Macromolecules 2001, 34, 7876-
7884.
26. Gao, C.; Yan, D.; Zhang, B.; Chen, W., Langmuir 2002, 18, 3708-3713.
27. Anghel, D. F.; Toca-Herrera, J. L.; Winnik, F. M.; Rettig, W.; Klitzing, R., Langmuir
2002, 18, 5600-5606.
28. Kanaganlingam, S.; Spartalis, J.; Cao, T.-M.; Duhamel, J., Macromolecules 2002,
35, 8571-8577.
Huy Tien Ngo Chapter 4
181
29. Siu, H.; Prazeres, T. J. V.; Duhamel, J.; Olesen, K.; Shay, G., Macromolecules 2005,
38, 2865-2875.
30. Siu, H.; Duhamel, J., Macromolecules 2005, 38, 7184-7186.
31. Ingratta, M.; Duhamel, J., Macromolecules 2007, 40, 6647-6657.
32. Seixas de Melo, J.; Costa, T.; Oliveira, N.; Schillén, K., Polym. Int. 2007, 56, 882-
899.
33. Seixas de Melo, J.; Costa, T.; Francisco, A.; Macanita, A. L.; Gago, S.; Goncalves, I.
S., Phys. Chem. Chem. Phys. 2007, 9, 1370-1385.
34. Siu, H.; Duhamel, J., J. Phys. Chem. B 2008, 112, 15301-15312.
35. Costa, T.; Schillén, K.; Miguel, M. G.; Lindman, B.; Seixas de Melo, J., J. Phys.
Chem. B 2009, 113, 6194-6204.
36. Hashidzume, A.; Ito, F.; Tomatsu, I.; Harada, A., Macromol. Rapid Commun. 2005,
26, 1151-1154.
37. Hashidzume, A.; Tomatsu, I.; Harada, A., Polymer 2006, 47, 6011-6027.
38. Costa, T.; Seixas de Melo, J., J. Polym. Sci. Pol. Chem. 2008, 46, 1402-1415.
39. Tomatsu, I.; Hashidzume, A.; Harada, A., J. Am. Chem. Soc. 2006, 128, 2226-2227.
40. Pouliquen, G.; Amiel, C.; Tribet, C., J. Phys. Chem. B 2007, 111, 5587-5595.
41. May, B. L.; Clements, P.; Tsanaktsidis, J.; Easton, C. J.; Lincoln, S. F., J. Chem. Soc,
Perkin Trans. 1 2000, 463-469.
42. Easton, C. J.; van Eyk, S. J.; Lincoln, S. F.; May, B. L.; Papageorgiou, J.; Williams,
M. L., Aust. J. Chem. 1997, 50, 9-12.
43. Pham, D.-T.; Ngo, H. T.; Lincoln, S. F.; May, B. L.; Easton, C. J., Tetrahedron 2010,
66, 2895-2898.
44. Lakowicz, J. R., Principles of Fluorescence Spectroscopy, 3rd Ed.; Springer
Science+Business Media: New York, 2006.
Huy Tien Ngo Chapter 4
182
45. Yang, C.; Mori, T.; Origane, Y.; Ko, Y. H.; Selvapalam, N.; Kim, K.; Inoue, Y., J.
Am. Chem. Soc. 2008, 130, 8574-8575.
Huy Tien Ngo Chapter 5 - Experimental
183
CHAPTER 5
EXPERIMENTAL†
† Publication associated with part of the material in this chapter:
Pham, D.-T.; Ngo, H. T.; Lincoln, S. F.; May, B. L.; Easton, C. J., Tetrahedron 2010, 66,
2895-2898.
Huy Tien Ngo Chapter 5 - Experimental
184
5.1. General
5.1.1. Instrumental
Routine 1D 1H and 13C NMR spectra were recorded on a Varian Gemini ACP-300
(300.145 MHz and 75.4 MHz, respectively) spectrometer, unless otherwise stated. Spectra
were obtained in either CDCl3, D2O or DMSO-d6 solutions with references to either
tetramethylsilane (H 0.0 for SiMe4) and CDCl3 (C 77.0) in CDCl3, the residual solvent
peak (H 2.49 and C 39.5) in DMSO-d6 or an external standard, aqueous
trimethylsilylpropiosulfonic acid, in D2O. Chemical shifts are cited on the scale in parts
per million, ppm, followed by multiplicity and assignment. The following abbreviations
are used to report multiplicity: s, single; d, doublet; t, triplet; q, quartet; m, multiplet; br,
broad. The value at the centre of the multiplet resonance is recorded excepted for signals
where a multiplet is well resolved in which case the values for all individual multiplet
components are given.
The 2D 1H ROESY and NOESY NMR spectra were recorded on a Varian Inova 600
(599.957 MHz) spectrometer, using a standard sequence with a mixing time of 300 ms.
Electrospray ionisation mass spectra, ESI-MS, were recorded on a Finnigan MAT ion
trap LC-Q octapole mass spectrometer. Gas chromatography - mass spectrometry, GC-MS,
data was obtained using a Shimadzu GC-MS spectrometer. Samples were dissolved in
either Milli-Q water, HPLC grade methanol or a mixture of both at a concentration of 0.5
mg cm-3. Elemental analyses were performed by the Microanalytical Service of the
Chemistry Department, University of Otago, New Zealand. Since cyclodextrin derivatives
contain associated water molecules, fractional numbers of water molecules were added to
the molecular formula to give the best fit to the microanalytical data.
Thin-layer chromatography, TLC, was carried out on Merck Kieselgel 60 F254 on
aluminium-backed sheets. For analysis of - and -cyclodextrin derivatives, plates were
developed with 7:7:5:4 v/v ethyl acetate/propan-2-ol/ammonium hydroxide/water. The
compounds were visualised by drying the plate, dipping it into a 1% sulphuric acid in
ethanol solution and followed by heating with a heat-gun. To visualise amino bearing
cyclodextrins, plates were dried prior to dipping into 0.5% ninhydrin in ethanol and heated
with a heat-gun before dipping in 1% sulphuric acid in ethanol. For the preparations of
Huy Tien Ngo Chapter 5 - Experimental
185
modified cyclodextrins described in the following sections, Rc represents the Rf of a
substituted cyclodextrin relative to the Rf of the parent cyclodextrin.
UV-visible absorbance spectra were recorded using a Varian CARY 5000 UV-VIS-NIR
spectrophotometer equipped with matched 1.0 cm path length quartz cells over a range of
required wavelengths at 0.5 nm intervals. Each solution was run against a reference
solution containing all components of the solution of interest except the absorbing
compound. Solutions were pre-equilibrated at 298.2 ± 0.2 K, unless stated otherwise and
maintained at this temperature during measurement by means of a thermostatted cell block.
All solutions were freshly prepared prior to measurement.
Fluorescence measurements were recorded using a Varian CARY Eclipse
spectrofluorimeter equipped with a 1.0 cm path length quartz cell. Spectra were obtained
over a range of desired wavelengths at 0.5 nm intervals, with both excitation and emission
slit widths of 5 nm (unless stated otherwise), using a 1% transmittance emission filter.
Emission spectra obtained were not corrected for instrumental factors. Solutions were pre-
equilibrated at 298.2 ± 0.2 K and maintained at this temperature during measurement by
means of a thermostatted cell block. All solutions were freshly prepared prior to
measurement.
Rheological measurements were carried out at the State Key Laboratory of Chemical
Engineering, East China University of Science and Technology, Shanghai 200237, China
using a Physica MCR 501 (Anton Parr GmbH) stress–controlled rheometer with a 25 mm
cone and plate geometry. Temperature was controlled at 298.2 ± 0.1 K by a Peltier plate.
Rheological samples were prepared by dissolution of PAA1NSen and PAA1NShn in 0.10
mol dm-3 aqueous sodium chloride to ensure screening of the electrostatic interactions
between the carboxylate groups. The solution pH was adjusted to 7.0 with 0.10 mol dm-3
aqueous sodium hydroxide solution.
5.1.2. Materials
All reagents were obtained from Sigma-Aldrich or other commercial sources and were
used without further purification, unless stated otherwise. Water was purified with a Milli-
Q system to give a resistivity of > 15 MΩ cm. Triethylamine (Ajax) was dried by
distillation. All organic solvents, N,N-dimethylformamide (APS), pyridine (Ajax), diethyl
ether (Chem Supply), acetone (Chem Supply), ethanol (Ajax), methanol (Ajax), N-
Huy Tien Ngo Chapter 5 - Experimental
186
methylpyrrolidin-2-one (Fluka), tetrahydrofuran (Chem Supply), dichloromethane (Chem
Supply), ethyl acetate (Chem Supply) were of HPLC grade and were used without further
purification.
Squat column chromatography was carried out using Merck Kieselgel 60 F254 thin layer
chromatography silica. Aluminium oxide column chromatography was carried out using
Acros Organics basic activated aluminium oxide, 50-200 micron, Brockman activity I with
appropriate amount of water added to give Brockman activity III. Bio-Rex 70 resin was
purchased from Bio-Rad Laboratories Inc., CA and was converted to the acid form using
3.0 mol dm-3 hydrochloric acid. Diaion HP-20 resin was purchased from Supelco, PA.
CD and CD were obtained from Nihon Shokuhin Kako Co. Unless otherwise stated,
6A-O-(4-methylbenzenesulfonyl)--cyclodextrin (6CDTs),1 6A-amino-6A-deoxy--
cyclodextrin (6CDNH2),2 were prepared according to literature methods. The modified
cyclodextrins were dried to a constant weight over P2O5 containing indicator (Sicapent)
under vacuum and stored in the dark under refrigeration. Pyronine B (PB+) was purchased
from Sigma as the 95% pure salt PB2Fe2Cl8, which was twice recrystallised from water
before use.3 The commercially obtained pyronine Y (PY+) chloride salt contained
approximately 40% impurities by weight. These water insoluble impurities were filtered
from an aqueous slurry with a 0.45 m filter before use.4 Hematoporphyrin (HP) was
purchased as the 95% pure dihydrochloride salt from Sigma and was used as received.
Poly(acrylic acid)s (PAA, Mw = 250,000, Mw/Mn ≈ 2) 35 wt% aqueous solution (Aldrich)
was diluted to approximately 10 wt% and freeze-dried to constant weight to give a white
solid. 1-Naphthalenesulfonyl chloride (Aldrich, 97%), succinyl chloride (Aldrich, 95%), 4-
nitrophenol (Sigma, 98%), 1,2-diaminoethane (Ajax), 1,6-diaminohexane (Aldrich) and
N,N’-dicyclohexylcarbodiimide (Merck, 98%) were used as supplied without further
purification.
5.1.3. Data Analysis
Equation 5.1 describes the observed absorbance when a single host-guest complex
equilibrium exists in the solution, where A, G, H.G represent the total absorbance, molar
absorbances of the guest and (host).(guest) complex, respectively. Equations 5.2–5.4
describe the observed absorbance when more than one equilibrium co-exist in the
Huy Tien Ngo Chapter 5 - Experimental
187
solutions, where G2, H.G2 represent the molar absorbances of the dimerised guest and 1:2
(host).(guest)2 complex, respectively.
A = G[guest] + H.G[(host).(guest)] (5.1)
A = G[guest] + G2[(guest)2] + H.G[(host).(guest)] (5.2)
A = G[guest] + H.G[(host).(guest)] + H.G2[(host).(guest)2] (5.3)
A = G[guest] + G2[(guest)2] + H.G[(host).(guest)] + H.G2[(host).(guest)2] (5.4)
The K1 for the 1:1 host-guest complexes of either PB+ or PY+ with CD and the linked
CD dimer hosts were derived by simultaneously fitting the algorithm analogous to Eqn.
5.1 to the absorbance variations over a wide wavelength range at 0.5 nm intervals, using
the non–linear least–squares SPECFIT/32 protocol.5 Analogous equations apply for the
fluorescence variations of all six systems.
The Kd for the dimerisation of HP2- was derived by simultaneously fitting the
algorithm analogous to Eqn. 5.2 to the absorbance variations over a wide wavelength
range at 0.5 nm intervals, with the absence of the third right–hand term, using the non–
linear least–squares fitting program HypSpec.6,7 Using the known values of Kd, HP and
HP2, the complexation constants, K1, for the 1:1 host/guest complex of HP2- by CD
and the linked CD dimer hosts were then derived by simultaneously fitting the
algorithm analogous to Eqn. 5.2 to the observed absorbance variations. Analogous
equations apply for the fluorescence variations of all three complex systems.
The stepwise complexation constants, K1 and K2, for 1:1 and 1:2 host–guest complexes
of either PAA1NSen or PAA1NShn with CD, CD, succindamide–linked CD dimers
and succinamide–linked CD dimers were derived by simultaneously fitting the algorithm
analogous to either Eqn. 5.1 or 5.3 to the fluorescence variations over a wide wavelength
range at 0.5 nm intervals, using the non–linear least–squares fitting program HypSpec.6,7
In the 1D 1H NMR study, the dimerisation constants, Kd, for PB+ and PY+ were derived
by simultaneously fitting dimerisation algorithm analogous to Eqn. 5.5 to the variation of
the 1H chemical shifts, exp, of the H1–H4 protons as [PB+]total and [PY+]total increased using
the HypNMR 2003 program.8,9 The K1 for the complexation of PB+ and PY+ by CD and
the linked CD dimer hosts were similarly derived by fitting Eqn. 5.6 to the 1H chemical
Huy Tien Ngo Chapter 5 - Experimental
188
shift variations, exp, of the PB+ and PY+ H1–H4 protons and either the PY+ H5 or PB+ H6
proton.
exp = PB[PB+] + PB2[(PB+)2] (5.5)
exp = PB[PB+] + PB2[(PB+)2] + CD.PB[CD.PB+] (5.6)
5.1.4. Molecular Modelling
Molecular modelling and MM2 energy minimisations were performed in the gas–phase
with the ChemBio3D® Ultra 11.0 software10 and geometry optimisation was performed
using the PM6 semi-empirical method in MOPAC2009.11,12 The Broyden-Fletcher-
Goldfarb-Shanno (BFGS) optimisation procedure was employed for all PM6
optimisation,13,14 and additional keywords, XYZ (for geometry optimisation using
Cartesian coordinates) and CHARGE=n were used as appropriate. The intitial molecular
models of the CD dimers: 33CD2suc and 66CD2suc and the hematoporphyrin
complexes: CD.HP2-, 33CD2suc.HP2- and 66CD2suc.HP2- in Chapter 3 were constructed
with the assistance of Dr. Duc-Truc Pham.
Huy Tien Ngo Chapter 5 - Experimental
189
5.2. Experimental for Chapter 2
5.2.1. Syntheses
2A-O-(4-Methylbenzenesulfonyl)--cyclodextrin, 2CDTs15
C2A O S
O
O
CD
The title compound was prepared according to the literature method,15 through the
reaction of CD (45.4 g, 0.04 mol) with p-toluenesulfonylchloride (28.8 g, 0.15 mol) in
DMF in the presence of dibutyltin oxide (25 g, 0.1 mol) and triethylamine (12.2 g, 0.12
mol) to afford a white powder with spectral information consistent with that in the
literature.16
Yield: 3.92 g (7.6 %)
TLC: Rc = 1.93
1H NMR: H (DMSO-d6) 7.86 (d, 2 H, Ar-H), 7.45 (d, 2 H, Ar-H), 5.69-5.91 (m, 13 H,
OH2,3), 4.82 (s, 7 H, H1), 4.48 (d, 7 H, OH6), 3.41-4.25 (m, 42 H, H2-6), 2.42 (s, 3 H, Ar-
CH3) 13C NMR: C (DMSO-d6): 133.4, 129.9, 128.2, 125.7 (Ar-C); 101.9-101.1 (C1B-G), 97.3
(C1A); 82.3-78.2 (C4); 73.1-69.2 (C2, C3, C5); 60.2 (C6); 40.9-38.4 (DMSO); 21.3 (Ar-
CH3).
2A,3A-Manno-epoxide--cyclodextrin, 23CDO17
C2A
O
C3ACD
The title compound was prepared by the literature method.17 A solution of 2A-O-(4-
methylbenzenesulfonyl)--cyclodextrin, 2CDTs (2.32 g, 1.8 mmol) in aqueous
ammonium bicarbonate (10 %, 100 cm3) was stirred at 60 oC for 3 hrs. The solvent was
Huy Tien Ngo Chapter 5 - Experimental
190
removed under vacuum and the residue was redissolved in water, followed by evaporation
to dryness (this procedure was repeated three times). This crude product was dissolved in
water (20 cm3) and added dropwise to vigorously stirred acetone (200 cm3). The precipitate
formed was collected by filtration and washed with acetone and diethyl ether to give a
crude product. The crude material was dissolved in water (100 cm3) and loaded onto a
Diaion HP-20 column (3 × 20 cm). The column was washed with water (500 cm3) and 10 –
20 % aqueous methanol and the washings were evaporated under vacuum to give the
product as a white powder, which contained traces of 2CDNH2 by-product. This solid was
run through a column (4.5 × 4.5 cm) of BioRex 70 (H+), 100-200 mesh (BioRad) and
eluted by water. The fractions containing the product were combined and evaporated to
dryness under vacuum to give the title compound as a white powder.
Yield: 1.45 g (72.1 %)
TLC: Rc = 1.11 1H NMR, H (D2O): 5.26 (s, 1 H, H1A-epoxide), 5.09-5.04 (m, 6 H, H1), 3.98-3.57 (m, 41
H, H2-6), 3.46 (d, 1 H, H2A-epoxide) 13C NMR, C(D2O): 104.5-103.7 (C1), 83.7-83.0 (C4), 75.7-72.1 (C2B-G, C3B-G, C5), 63.6
(C6B-G), 62.9 (C6A), 57.1 (C2A), 52.2 (C3A).
3A-Amino-3A-deoxy-(2AS,3AS)--cyclodextrin, 3CDNH218
NH2C3A
CD
The title compound was prepared by the literature method.18 2A,3A-Manno-epoxide--
cyclodextrin (1.45 g, 1.3 mmol) was dissolved in aqueous ammonium hydroxide (25%, 40
cm3) and the solution was stirred at 60 oC for 4 hrs. The mixture was then evaporated to
dryness and the residual was dissolved in aqueous ammonium hydroxide (28%, 10 cm3)
and added to acetone (200 cm3). The precipitate was collected, washed with acetone and
diethyl ether and dried under vacuum to a crude product. The solid was dissolved in water
(10 cm3) and loaded onto a column (4.5 × 4.5 cm) of BioRex 70 (H+), 100-200 mesh
(BioRad). After flushing with water (ca. 400 cm3), the 3CDNH2 product was eluted with
1.0 mol dm-3 aqueous ammonium hydroxide (ca. 100 cm3 fractions). Fractions containing
the product were combined and evaporated to dryness under vacuum (removal of excess
Huy Tien Ngo Chapter 5 - Experimental
191
ammonia was achieved by dissolving the residue in water and evaporating to dryness three
times) to afford 3CDNH2 as a white powder.
Yield: 0.88 g (60%)
TLC: Rc = 0.71 1H NMR: H (D2O): 5.13 (d, 2 H, H1A), 5.04-4.93 (m, 5 H, H1), 4.23 (m, 1 H, H2A), 4.01-
3.54 (m, 40 H, H2-6), 2.98 (d, 1 H, H3A) 13C NMR: C (D2O): 103.1-99.7 (C1), 80.9-80.2 (C4), 79.2-71.1 (C2, C3B-G, C5), 60.8-
59.8 (C6), 52.2 (C3A).
Bis(4-nitrophenyl) succinate19,20
O
O
O
O
NO2
O2N
A method silimar to those reported in the literature was used to prepared the title
compound.19,20 To a stirred solution of succinyl chloride (4.83 g, 31.2 mmol) in
dichloromethane (100 cm3) was added 4-nitrophenol (9.87 g, 71 mmol) in one portion and
then triethylamine (7.54 g, 74.5 mmol) dropwise over a period of 15 mins. The mixture
was stirred for 1 hr at room temperature. The reaction mixture was subsequently washed
with water (2 x 100 cm3) and the organic layer was dried over anhydrous MgSO4. TLC
(10% hexane in dichloromethane) of the organic extract showed the title compound (Rf =
0.6) and 4-nitrophenol (Rf = 0.2). Evaporation of the solvent yielded the crude product
which was recrystallised from ethyl acetate to afford the pure product as a cream coloured
flaky powder.
Yield: 2.04 g (18.1 %) 1H NMR: H (CDCl3) 8.27 (d, 4 H, ArH), 7.29 (d, 4 H, ArH), 3.06 (s, 4 H, CH2) 13C NMR: C (CDCl3) 166.8 (ester C=O), 155-122.2 (ArC), 29.1 (succinyl CH2).
General procedure for the preparation of the succinamide-linked CD dimers21
The succinmamide-linked CD dimers were prepared according to the literature
method.21 Either (2AS,3AS)-3A-amino-3A-deoxy--cyclodextrin or 6A-amino-6A-deoxy--
cyclodextrin (~1 mmol) was dissolved in pyridine (20 cm3) and stirred at room temperature
Huy Tien Ngo Chapter 5 - Experimental
192
for 15 min. Bis(4-nitrophenyl) succinate (0.4 equivalents) was added to this solution in two
or more portions over a period of 1 hr. The reaction mixture was then stirred for 48 hrs at
room temperature before being added dropwise to diethyl ether (200 cm3) with vigorous
stirring. The resultant precipitate was collected by centrifugation, washed with acetone and
diethyl ether and dried under vacuum. The product was dissolved in H2O and run down a
BioRex 70 (H+) column to remove either excess (2AS,3AS)-3A-amino-3A-deoxy--
cyclodextrin or 6A-amino-6A-deoxy--cyclodextrin. The white solid products were
obtained by freeze drying followed by further drying over phosphorous pentoxide.
N,N′-Bis((2AS,3AS)-3A-deoxy--cyclodextrin-3A-yl) succinamide, 33CD2suc
HNC3A
O
HN C3A
OCD CD
The title compound was prepared by treatment of the 3CDNH2 (880 mg, 0.78 mmol) with
bis(4-nitrophenyl) succinate (112.7 mg, 0.31 mmol) according to the general procedure.
After the general work-up and purification procedure, the title compound was obtained as a
white solid.
Yield: 0.81 g (88%)
TLC: Rc = 0.54 1H NMR: H (D2O): 5.11-4.92 (m, 14 H, H1), 4.21-3.56 (m, 84 H, H2-6), 2.6 (s, 4 H, CH2) 13C NMR, C (D2O): 177.6 (C=O), 106.5, 104.6, 104.5, 104.1, 103.9 (C1), 83.8, 83.6, 83.5,
83.3, 82.7 (C4), 75.9, 75.7, 75.3, 75.0, 74.8, 74.5, 74.2, 74.0, 72.6 (C2,3,5), 63.0, 62.4,
(C6), 53.7 (C3A), 33.7 (CH2)
Mass spectrum m/z: 2350 (M+), 2373 (M + Na)+
Elemental analysis: C88H144N2O70.20H2O: C, 38.99; H, 6.84; N, 1.03. Found: C, 39.1; H,
6.5; N, 1.1.
Huy Tien Ngo Chapter 5 - Experimental
193
N,N′-Bis(6A-deoxy--cyclodextrin-6A-yl) succinamide, 66CD2suc
HN
O
C6AHN
O
C6A
CD CD
The title compound was prepared by treatment of the 6CDNH2 (1.21 g, 1.1 mmol) with
bis(4-nitrophenyl) succinate (158.5 mg, 0.44 mmol) according to the general procedure.
After the general work-up and purification procedure, the title compound was obtained as a
white solid.
Yield: 0.84 g (81.2 %)
TLC: Rc = 0.40 1H NMR, H (D2O): 5.07 (m, 14 H, H1), 3.99-3.38 (m, 84 H, H2-H6), 2.60 (m, 4 H, CH2)
13C NMR, C (D2O): 177.5 (C=O), 104.6 (C1), 85.7, 83.8 (C4), 75.8, 74.8, 74.5, 72.9
(C2,3,5), 63.0 (C6B-G), 42.8 (C6A), 33.7 (CH2)
Mass spectrum m/z: 2351 (M + H)+, 2373 (M+Na)+
Elemental analysis: C88H144N2O70.19H2O: C, 39.25; H, 6.81; N, 1.04. Found: C, 39.3; H,
6.6; N, 1.1.
5.2.2. Sample Preparation
5.2.2.1. Sample Preparation for UV–vis and Fluorescence Studies
Stock solutions of the pyronines PB+ (6.0 × 10-4 mol dm-3), PY+ (9.0 × 10-4 mol dm-3)
and the hosts CD (1.5 × 10-2 mol dm-3), 33CD2suc and 66CD2suc (5.0 × 10-3 mol dm-3)
were freshly prepared in aqueous hydrochloric acid (1.00 × 10-4 mol dm-3),22 to prevent
base hydrolysis and 0.10 mol dm-3 sodium chloride to maintain constant ionic strength.
The concentrations of PY+ stock were estimated using the reported molar absorptivity at
546 nm of = 8.1 × 104 mol-1 dm3 cm-1.23
Aqueous solutions for UV–vis and fluorescence studies were 6.0 × 10-6 mol dm-3 in PB+
and 9.0 × 10-6 mol dm-3 in PY+; and 6.0 x 10-7 mol dm-3 of PB+ and 9.0 × 10-7 mol dm-3 of
PY+, respectively. The CD and linked CD dimer concentrations varied over wide ranges,
as indicated in the figure captions.
Huy Tien Ngo Chapter 5 - Experimental
194
5.2.2.2. Sample Preparation for 1H NMR Studies
Solutions for 1H NMR experiments were prepared in D2O, 1.00 × 10-4 mol dm-3 in
hydrochloric acid and 0.10 mol dm-3 in sodium chloride. The concentrations of PB+ and
PY+ solutions for dimerisation studies ranged from 1.0 × 10-3 mol dm-3 to 2.0 × 10-2 mol
dm-3. For complexation studies, the concentrations of PB+ and PY+ were kept constant at
2.0 × 10-3 mol dm-3, while those of the CD and CD dimer hosts were varied from 0–5.0
× 10-3 mol dm-3 by addition of appropriate volumes of stock solutions.
For the 2D 1H ROESY NMR experiments, each sample was 2.0 × 10-3 mol dm-3 in
either PB+ or PY+ and in either CD or a linked CD dimer, unless stated otherwise in the
figure captions.
Huy Tien Ngo Chapter 5 - Experimental
195
5.3. Experimental for Chapter 3
5.3.1. Syntheses24
6A-O-(2,4,6-triisopropylbenzenesulfonyl)--cyclodextrin (6CDTPBS)25
SO
OO
CDC6A
The title compound was prepared according to the literature method.25 To a solution
pyridine (30 cm3) was added CD (5.0 g, 3.85 mmol) and the mixture was stirred at room
temperature for 30 mins under nitrogen. 2,4,6-triisopropylbenzenesulfonyl chloride (3.5 g,
11.56 mmol) was added in 3 portions over the period of 2 hrs and stirred for 24 hrs. The
mixture was concentrated to ca. 10 cm3 and added dropwise to vigorously stirred acetone
(200 cm3). The resulting precipitate was collected by filtration, washed with acetone and
diethyl ether and dried under vacuum to give 6.4 g of crude product. The crude material
was repeatedly recrystallised from water until no more crystals formed to afford the title
product as a white solid.
Yield: 0.78 g (12.9 %)
TLC: Rc = 1.32
1H NMR: H (DMSO-d6) 7.28 (s, 2 H, Ar-H), 5.91-5.69 (m, 16 H, OH2,3), 4.85 (m, 8 H,
H1), 4.58-4.47 (m, 7 H, OH6), 4.25 (m, 2 H, CH2-OSO-Ar), 3.21-4.03 (m, 47 H, H2-6, Ar-
CH(CH3)2), 2.94 (m, 1 H, Ar-CH(CH3)2), 1.21 (m, 18 H, Ar-CH(CH3)2).
2A-O-(4-methylbenzenesulfonyl)--cyclodextrin, 2CDTs, and its 6A analogue, 6CDTs
C6AC2A OO S
O
O
S
O
O
CDCD
2CDTS 6CDTS
Huy Tien Ngo Chapter 5 - Experimental
196
The title compounds were prepared by a modification of the literature method.15 CD
(51.9 g, 40.0 mmol) was added to anhydrous DMF (150 cm3) and the mixture was stirred
for two hrs at 0 oC under dry nitrogen until dissolution was complete. After heating to 100 oC, dibutyltin oxide (25.1 g, 100.9 mmol) was added and stirred for another 2 hrs. The
mixture was then cooled to 0 oC, triethylamine (12.2 g, 120.6 mmol) was added, followed
by dropwise addition of 4-toluenesulfonyl chloride (20 g, 105 mmol) in DMF (50 cm3).
The mixture was stirred for 2 hrs before another portion of 4-toluenesulfonyl chloride (9.7
g, 50.9 mmol) in DMF (20 cm3) was added dropwise. The resultant solution was stirred for
a further 10 hrs at room temperature and then concentrated to a yellow syrup. This was
added to 2 dm3 of vigorously stirred acetone and stirring was continued for 30 mins. The
precipitate formed was collected by filtration, washed with acetone and diethyl ether and
dried under vacuum to give 62 g of crude product which was recrystallised from ca. 200
cm3 water. The precipitate was collected and dried under vacuum to give ca. 9 g of crude
6CDTs, while the filtrate was evaporated to dryness to give ca. 47 g of crude 2CDTs.
The crude 2CDTs was dissolved in water (1 dm3) and loaded onto a Diaion HP-20
column (5 × 30 cm). After flushing with ca. 3 dm3 of water, followed by 10 – 15 %
aqueous methanol solvent gradient elution of unreacted CD, 2CDTs was eluted with 20 -
25% aqueous methanol (ca. 400 cm3 fractions). The fractions containing the product were
combined, the methanol was removed and the product was dried under vacuum to give the
2CDTs as a white powder.
Yield: 4.37 g (7.5 %)
TLC: Rc = 1.67 1H NMR: H (DMSO-d6): 7.83, 7.47 (ABq, J = 8.2 Hz, 4H, ArH), 5.91-5.69 (m, 15 H,
OH2, OH3), 4.88 (s, 8 H, H1), 4.30-3.30 (m, 56 H, H2-6, OH6), 2.41 (s, 3 H, Ar-CH3) 13C NMR: C (DMSO-d6): 133.4, 129.9, 128.2, 125.7 (Ar-C); 101.9-101.1 (C1B-H), 97.3
(C1A); 82.3-78.2 (C4); 73.1-69.2 (C2, C3, C5); 60.2 (C6); 40.9-38.4 (DMSO); 21.3 (Ar-
CH3).
The crude 6CDTs was dissolved in water (500 – 700 cm3) and loaded onto a Diaion
HP-20 column (3 × 25 cm). After flushing with water (ca. 1 dm3), followed by 10 – 20%
aqueous methanol solvent gradient elution of unreacted CD, 6CDTs was eluted with 30 –
Huy Tien Ngo Chapter 5 - Experimental
197
40 % aqueous methanol (ca. 250 cm3 fractions). The fractions containing the product were
combined and evaporated to dryness under vacuum to give 6CDTs as a white powder.
Yield: 2.47 g (4.25 %)
TLC: Rc = 1.40 1H NMR: H (DMSO-d6): 7.78, 7.46 (ABq, J = 8.3 Hz, 4H, ArH), 6.05-5.35 (m, 16 H,
OH2, OH3), 5.09-4.81 (m, 8 H, H1), 4.31-3.20 (m, 55 H, H2-6, OH6), 2.42 (s, 3 H, Ar-CH3) 13C NMR: C (DMSO-d6): 133.2, 130.7, 128.8, 126.2 (Ar-C); 102.9-101.8 (C1A); 81.6-80.8
(C4); 73.6-69.7 (C2, C3, C5); 60.7 (C6); 40.9-38.4 (DMSO); 21.8 (Ar-CH3).
2A,3A-Manno-epoxide--cyclodextrin, 23CDO
C2A
O
C3ACD
The title compound was prepared according to the literature method.17 A solution of 2A-
O-(4-methylbenzenesulfonyl)--cyclodextrin, 2CDTs (4 g, 2.76 mmol) in aqueous
ammonium bicarbonate (10 %, 125 cm3) was stirred at 60 oC for 3 hrs. The solvent was
removed under vacuum and the residue was redissolved in water, followed by evaporation
to dryness (this procedure was repeated three times). This crude product was dissolved in
water (20 cm3) and added dropwise to vigorously stirred acetone (500 cm3). The precipitate
formed was collected by filtration and washed with acetone and diethyl ether to give 4 g of
crude product. The crude material was dissolved in water (125 cm3) and loaded onto a
Diaion HP-20 column (3 × 20 cm). The column was washed with water (1 dm3) and 10 %
aqueous methanol and the washings were evaporated under vacuum to give the product as
a white powder, which contained traces of 2CDNH2 by-product. This solid was run
through a column (4.5 × 4.5 cm) of BioRex 70 (H+), 100-200 mesh (BioRad) and eluted by
water. The fractions containing the product were combined and evaporated to dryness
under vacuum to give the title compound as a white powder.
Yield: 2.77 g (78.6 %)
TLC: Rc = 1.15 1H NMR: H (D2O): 5.27 (s, 1 H, H1A-epoxide), 5.13-5.07 (m, 7 H, H1), 3.95-3.59 (m, 47
H, H2-6), 3.49 (d, 1 H, H2A-epoxide)
Huy Tien Ngo Chapter 5 - Experimental
198
13C NMR: C(D2O): 104.5-103.7 (C1), 83.7-83.0 (C4), 75.7-72.1 (C2B-H, C3B-H, C5), 63.6
(C6B-H), 62.9 (C6A), 57.1 (C2A), 52.2 (C3A).
3A-Amino-3A-deoxy-(2AS,3AS)--cyclodextrin, 3CDNH2
NH2C3A
CD
The title compound was prepared according to the literature method.18 2A,3A-Manno-
epoxide--cyclodextrin, 23CDO (2.6 g, 2.03 mmol) was dissolved in aqueous ammonium
hydroxide (25%, 60 cm3) and the solution was stirred at 60 oC for 4 hrs. The mixture was
then evaporated to dryness and the residual was dissolved in aqueous ammonium
hydroxide (28%, 20 cm3) and added to acetone (500 cm3). The precipitate was collected,
washed with acetone and diethyl ether and dried under vacuum to obtain 2.74 g of the
crude product. This was dissolved in water (20 cm3) and loaded onto a column (4.5 × 4.5
cm) of BioRex 70 (H+), 100-200 mesh (BioRad). After flushing with water (ca. 500 cm3),
the 3CDNH2 product was eluted with 1.0 mol dm-3 aqueous ammonium hydroxide (ca.
100 cm3 fractions). Fractions containing the product were combined and evaporated to
dryness under vacuum (removal of excess ammonia was achieved by dissolving the residue
in water and evaporating to dryness three times) to afford 3CDNH2 as a white powder.
Yield: 1.49 g (56.7 %)
TLC: Rc = 0.76. 1H NMR, H (D2O): 5.22 (d, 2 H, H1A), 5.16-4.93 (m, 6 H, H1), 4.20 (m, 1
H, H2A): 4.00-3.56 (m, 46 H, H2-6), 3.10 (d, 1 H, H3A) 13C NMR: C (D2O): 103.1-99.7 (C1), 80.9-80.2 (C4), 79.2-71.1 (C2, C3B-H, C5), 60.8-
59.8 (C6), 52.2 (C3A).
6A-Amino-6A-deoxy--cyclodextrin, 6CDNH2
NH2C6A
CD
Huy Tien Ngo Chapter 5 - Experimental
199
The title compound was prepared according to the literature method.2 6A-O-(4-
Methylbenzenesulfonyl)--cyclodextrin, 6CDTs (3.4 g, 2.3 mmol) was dissolved in
ammonium hydroxide (28%, 250 cm3) at 0 oC. The reaction vessel was closed and left in
the dark with occasional stirring for 5 days. The ammonium hydroxide was removed under
reduced pressure, after which water was added (100 cm3) and removed under reduced
pressure. The remaining solid was dissolved in ammonium hydroxide (28%, 20 cm3) and
the solution added drop-wise to vigorously stirring acetone (450 cm3) and stirred for 30
min. The resulting precipitate was dried under vacuum to give crude 6CDNH2 as a cream
powder. This was dissolved in water (20 cm3) and loaded onto a BioRex 70 (H+) column.
The column was washed with water (400 cm3), and the title compound was eluted with
0.05 – 0.1 mol dm-3 aqueous ammonium carbonate (4 × 100 cm3 fractions). The 6CDNH2
containing fractions were evaporated to dryness under reduced pressure, and the residue
freeze dried to give 6CDNH2 as a white solid.
Yield: 0.55 g (18 %)
TLC: Rc = 0.90 1H NMR: δ(D2O) 5.37 (s, 2H, H1A), 5.10 (m, 14H, H1B-H), 3.93-3.58 (m, 48H, H2-6) 13C NMR: δ(D2O): 103.72-101.77 (C1); 80.50-79.0 (C4); 73.89-71.88 (C2, C3, C5); 60.33
(C6); 41.05 (C6A).
General procedure for the preparation of the succinamide-linked CD dimers21
The succinmamide-linked CD dimers were prepared according to the literature
method.21 Either (2AS,3AS)-3A-amino-3A-deoxy--cyclodextrin or 6A-amino-6A-deoxy--
cyclodextrin (~1 mmol) was dissolved in pyridine (20 cm3) and stirred at room temperature
for 15 min. Bis(4-nitrophenyl) succinate (0.4 equivalents) was added to this solution in two
or more portions over a period of 1 hr. The reaction mixture was then stirred for 48 hrs at
room temperature before being added dropwise to diethylether (200 cm3) with vigorous
stirring. The resultant precipitate was collected by centrifugation, washed with acetone and
diethylether and dried under vacuum. The product was dissolved in H2O and run down a
BioRex 70 (H+) column to remove either excess (2AS,3AS)-3A-amino-3A-deoxy--
cyclodextrin or 6A-amino-6A-deoxy--cyclodextrin. The white solid products were
obtained by freeze drying followed by further drying over phosphorous pentoxide.
Huy Tien Ngo Chapter 5 - Experimental
200
N,N′-Bis((2AS,3AS)-3A-deoxy--cyclodextrin-3A-yl) succinamide, 33CD2suc
HNC3A
O
HN C3A
OCD CD
The title compound was prepared by treatment of the 3CDNH2 (1.39 g, 1.08 mmol)
with bis(4-nitrophenyl) succinate (155 mg, 0.43 mmol) according to the general procedure.
After the general work-up and purification procedure, the title compound was obtained as a
white solid.
Yield: 0.87 g (75.7 %)
TLC: Rc = 0.60 1H NMR: H (D2O): 5.39-4.94 (m, 16 H, H1), 4.26-3.59 (m, 96 H, H2-6), 2.6 (s, 4 H, CH2) 13C NMR: C (D2O): 177.7 (C=O), 105.8, 104.2, 102.3 (C1), 81.9, 82.7, 83.1 (C4), 72.3,
73.9, 74.4, 74.9, 74.9, 75.2, 75.6 (C2,3,5), 62.4, 62.9 (C6), 53.6 (C3A), 33.6 (CH2)
Mass spectrum m/z: 1359.9 (M+Na)2+
Elemental analysis: C100H164N2O80.19H2O: C, 39.81; H, 6.75; N, 0.93. Found: C, 39.54; H,
6.39; N, 0.89.
N,N′-Bis(6A-deoxy--cyclodextrin-6A-yl) succinamide, 66CD2suc
HN
O
C6AHN
O
C6A
CD CD
The title compound was prepared by treatment of the 6CDNH2 (1.58 g, 1.21 mmol)
with bis(4-nitrophenyl) succinate (176 mg, 0.49 mmol) according to the general procedure.
After the general work-up and purification procedure, the title compound was obtained as a
white solid.
Yield: 1.2 g (91.6 %)
TLC: Rc = 0.45 1H NMR, H (D2O): 5.11 (m, 16 H, H1), 3.42-3.93 (m, 96 H, H2-H6), 2.59 (m, 4 H, CH2)
Huy Tien Ngo Chapter 5 - Experimental
201
13C NMR, C (D2O): 177.5 (C=O), 104.3 (C1), 85.7, 83.1 (C4), 75.6, 75.0, 74.4, 72.8
(C2,3,5), 62.9 (C6B-H), 42.8 (C6A), 33.7 (CH2)
Mass spectrum m/z: 1359.9 (M+Na)2+
Elemental analysis: C100H164N2O80.20H2O: C, 39.58; H, 6.78; N, 0.92. Found: C, 39.48; H,
6.40; N, 0.91.
5.3.2. Sample Preparation
5.3.2.1. Sample Preparation for UV–vis and Fluorescence Studies
Solutions were prepared from fresh stock solutions in pH 10.0, 0.025 mol dm-3
carbonate/bicarbonate buffer (NaHCO3 0.0107 mol dm-3, Na2CO3 0.0143 mol dm-3 and
NaCl 0.0466 mol dm-3) at constant ionic strength I = 0.10 mol dm-3. Initial aqueous
solutions for UV–visible studies were 7.6 × 10-6 mol dm-3 in HP2- and the titrations
were conducted by sequential injection of 0.100 cm3 aliquots of either the buffer (in
dimerisation studies) or CD (2.63 × 10-2 mol dm-3) and linked CD dimers (2.50 × 10-3
mol dm-3) in complexation studies.
Solutions for fluorescence studies were 2.10 × 10 -7 mol dm-3 in HP2- and the CD
and linked CD dimer concentrations varied over wide ranges as indicated in the figure
captions.
5.3.2.2. Sample Preparation for 1H NMR Studies
Solutions for 2D 1H–NOESY NMR experiments were prepared in D2O (pD 10.0
carbonate/bicarbonate buffer, NaHCO3 0.0107 mol dm-3, Na2CO3 0.0143 mol dm-3 and
NaCl 0.0466 mol dm-3) at constant ionic strength I = 0.10 mol dm-3. Each sample was
5.0 × 10-3 mol dm-3 in HP2- and equimolar in either CD or a linked CD dimer.
5.3.3. Thermodynamic Parameters Determination
UV–vis titrations at temperatures ranging from 278.2 to 318.2 K were performed to
obtain the dimerisation constants, Kd, of HP2- and the complexation constants, K1, of
the 1:1 host-guest complexes.
The relationship between the Gibbs free energy (Go), enthalpy (Ho) and entropy
(So) for the dimerisation or complexation and the equilibrium constants (K) are given
Huy Tien Ngo Chapter 5 - Experimental
202
by the following van’t Hoff equations:
Go = –RTlnK (5.6)
with
Go = Ho – TSo (5.7)
from 5.6 and 5.7,
lnK = –Ho/RT + So/R (5.8)
where R is the gas constant and T is the absolute temperature. The plot of lnK versus 1/T
according to Eqn. 5.8 is a van’t Hoff plot, and for a linear plot, the slope and the intercept
represent –Ho/R and So/R, respectively.
Huy Tien Ngo Chapter 5 - Experimental
203
5.4. Experimental for Chapter 4
5.4.1. Syntheses
4-Nitrophenyl naphthalene-1-sulfonate
SO
OO N+
O
O-
This procedure was adopted from a similar reported method.26 A mixture of 4-
nitrophenol (2.38 g, 17.1 mmol), 1-naphthalenesulfonyl chloride (3.88 g, 17.1 mmol) and
triethylamine (2.60 g, 25.6 mmol) in dichloromethane (200 cm3) was stirred at room
temperature for 3 hrs under nitrogen. The reaction mixture was filtered through Celite and
the filtrate was loaded onto a squat column (4.5 × 9 cm) and eluted with dichloromethane
(500 cm3). Fractions containing the product was combined and evaporated under reduced
pressure to give the pure product as an orange powder.
Yield: 5.09 g (90.4 %)
TLC (10 % hexane in CH2Cl2): Rf = 0.73 1H NMR: H (DMSO-d6): 8.64 (d, J = 8.4 Hz, 1H, naphthyl H8), 8.46 (d, J = 8.4 Hz, 1H,
naphthyl H2), 8.25-8.16 (m, 4H, naphthyl H4,5, phenyl H3,4), 7.96 (t, J = 6.9 Hz, 1H,
naphthyl H7), 7.82 (t, J = 6.9 Hz, 1H, naphthyl H6), 7.66 (t, J = 6.9 Hz, 1H, naphthyl H3),
7.23-7.17 (m, 2H, phenyl H2,6).
N-(2-Aminoethyl)-1-naphthyl-sulfonamide, 1NSen
SO O
HN
NH2
A solution of 4-nitrophenyl naphthalene-1-sulfonate (600 mg, 1.82 mmol) in
dichloromethane was added dropwise to a vigorously stirred solution of 1,2-diaminoethane
(1.1 g, 18.2 mmol) in dichloromethane (10 cm3) at room temperature over a period of 4
hrs. The reaction was left overnight before being diluted with water (50 cm3) and acidified
Huy Tien Ngo Chapter 5 - Experimental
204
to pH < 2 with 3.0 mol dm-3 hydrochloric acid. The solution was washed with
dichloromethane (3 × 50 cm3), the aqueous solution was made basic to pH > 10 with 40
wt% aqueous sodium hydroxide and extracted with dichloromethane (3 × 50 cm3). The
combined dichloromethane solution was washed successively with water (3 × 100 cm3)
and brine (3 × 50 cm3) and dried over anhydrous sodium sulphate. The solvent was
removed under reduced pressure to give the pure product as a slight yellow solid.
Yield: 262 mg (57.5 %) 1H NMR: H (DMSO-d6): 8.66 (d, J = 8.4 Hz, 1H, ArH8), 8.22 (d, J = 8.4 Hz, 1H, ArH2),
8.11 (m, 2H, ArH4,5), 7.74-7.62 (m, 3H, ArH3,6,7), 2.75 (t, J = 6.6 Hz, 2H, SO2NHCH2),
2.43 (t, J = 6.6 Hz, 2H, CH2NH2).
GC-MS: C12H14N2O2S m/z calcd. 250.32, found: 250.15.
N-(6-Aminohexyl)-1-naphthyl-sulfonamide, 1NShn
SO OHN
NH2
A solution of 4-nitrophenyl naphthalene-1-sulfonate (585 mg, 1.78 mmol) in N,N-
dimethylformamide (10 cm3) was added dropwise to a vigorously stirred solution of 1,6-
diaminohexane (2.25 g, 19.4 mmol) in N,N-dimethylformamide (10 cm3) at room
temperature over a period of 4 hrs and the reaction was left overnight. The solvent was
removed under reduced pressure and dissolved in dichloromethane (40 cm3) and 10 wt%
hydrochloric acid (40 cm3). The aqueous solution was washed with dichloromethane (3 ×
40 cm3), made basic to pH > 10 with 40 wt% aqueous sodium hydroxide and extracted
with dichloromethane (3 × 50 cm3). The combined dichloromethane solution was washed
successively with water (3 × 50 cm3) and brine (3 × 50 cm3) and dried over anhydrous
sodium sulphate. The solvent was concentrated down to approximately 5 cm3, any
precipitation formed was filtered and the filtrate was loaded onto a basic aluminium oxide
column (4.5 × 4.5 cm, Brockman activity III). The column was eluted with a gradient of
dichloromethane:methanol from 100:0 to 70:30. Fractions containing the product were
combined and concentrated under reduced pressure to give the product as yellow oil.
Yield: 176 mg (32 %)
Huy Tien Ngo Chapter 5 - Experimental
205
1H NMR: H (DMSO-d6): 8.65 (d, J = 7.8 Hz, 1H, ArH8), 8.22 (d, J = 8.1 Hz, 1H, ArH2),
8.10 (m, 2H, ArH4,5), 7.71-7.62 (m, 3H, ArH3,6,7), 2.76 (t, J = 6.9 Hz, 2H, SO2NHCH2),
2.36 (t, J = 6.9 Hz, 2H, CH2NH2), 1.27-1.02 (m, 8H, (CH2)4)
GC-MS: C16H22N2O2S m/z calcd. 306.42, found: 306.20.
General procedure for the preparation of the 3% randomly substituted 1-naphthyl-
sulfonamide poly(acrylate)s27,28
The 3% randomly substituted 1-naphthyl-sulfonamide poly(acrylate)s were prepared
according to the generam procedure reported in the literature.27,28 The solid poly(acrylic
acid)s, PAA (1.9 g, 26.4 mmol of -COOH groups) was dissolved in N-methylpyrrolidin-2-
one (60 cm3) at 60 oC for 24 hrs. Either 1NSen or 1NShn (0.79 mmol) in N-
methylpyrrolidin-2-one (7.5 cm3) was added followed by dicyclohexylcarbodiimide (0.79
mmol) in N-methylpyrrolidin-2-one (7.5 cm3) and the reaction mixture was stirred at 60 oC
for at least 48 hrs. After cooling to room temperature, 40 wt% aqueous sodium hydroxide
(60 cm3) was added. The resulting precipitate was filtered and washed with 60 oC N-
methylpyrrolidin-2-one (2 × 30 cm3) and methanol (2 × 40 cm3). The crude product was
dissolved in water (12.5 cm3) and added dropwise to methanol (100 cm3) and the
precipitate collected (this step was repeated). The solid was dissolved in water (30 cm3)
and dialysed (Spectra/Por 3 tubing, molecular weight cutoff 3,500 g mol-1) against
deionised water until the conductivity of the water outside the tube remained constant. The
final dry product was obtained as the sodium polyarylate salt by freeze-drying after
concentrating the solution to 10 cm3 by evaporation. Typical yields were 80–90 %. The
degree of substitution was determined to be 3.0 ± 0.3 % by 1H NMR spectroscopy
according to the literature method.27,28 The 1H NMR spectra of PAA1NSen and
PAA1NShn in D2O are shown in Chapter 4.
5.4.2. Sample Preparation
5.4.2.1.Sample Preparation for UV–vis and Fluorescence Studies
Stock solutions of 5.0 wt% in either PAA1NSen or PAA1NShn were prepared in pH
7.0 phosphate buffer (KH2PO4 0.0195 mol dm-3, Na2HPO4 0.0268 mol dm-3) at constant
ionic strength I = 0.10 mol dm-3. For UV–vis and fluorescence studies, the stock
solutions were diluted to make 0.0330 wt% and 0.0033 wt% in either PAA1NSen or
PAA1NShn, which correspond to 1.0 × 10-4 mol dm-3 and 1.0 × 10-5 mol dm-3 in either
Huy Tien Ngo Chapter 5 - Experimental
206
1NSen or 1NShn groups, respectively, based on their calculated molecular weights
determined below.
Determination of the molecular weights of 1NSen and 1NShn in 3% randomly
substituted PAAs:
Mw(1NSen) = 3 × [Mw(CH2CHCOONa) × 97 % + Mw(CH2CHCO1NSen) × 3 %]
= 3343 g mol-1
Mw(1NShn) = 3 × [Mw(CH2CHCOONa) × 97 % + Mw(CH2CHCO1NShn) × 3 %]
= 3399 g mol-1
The initial solutions for fluorescence studies of 1.0 × 10-5 mol dm-3 in either [1Nsen]
or [1NShn] were sequentially diluted with 0.050 cm3 aliquots of stock solutions of each
CD host: CD (1.06 × 10-2 mol dm-3), CD (4.96 × 10-2 mol dm-3), 33CD2suc (2.49 ×
10-3 mol dm-3), 66CD2suc (2.31 × 10-3 mol dm-3), 33CD2suc (2.63 × 10-3 mol dm-3) or
66gCD2suc (2.49 × 10-3 mol dm-3). In a typical titration measurement, 30 aliquots of
either host were added.
5.4.2.2. Sample Preparation for 1H NMR Studies
Solutions for 1D and 2D 1H–NOESY NMR experiments were prepared in D2O in
0.10 mol cm-3 sodium chloride. The solution pH was adjusted to 7.0 with 0.10 mol dm -3
aqueous sodium hydroxide solution. Each sample contained 1.43 wt% in either
PAA1NSen or PAA1NShn (3.0 × 10-3 mol dm-3 in 1NSen groups and 2.94 × 10-3 mol
dm-3 in 1NShn groups) and the same concentration of CD, CD or a linked CD dimer.
Huy Tien Ngo Chapter 5 - Experimental
207
5.5. References
1. Brady, B.; Lynam, N.; O'Sullivan, T.; Ahern, C.; Darcy, R., Org. Syntheses 2000,
77, 220-224.
2. Brown, S. E.; Coates, J. H.; Coghlan, D. R.; Easton, C. J.; Vaneyk, S. J.;
Janowski, W.; Lepore, A.; Lincoln, S. F.; Luo, Y.; May, B. L.; Schiesser, D. S.;
Wang, P.; Williams, M. L., Aust. J. Chem. 1993, 46, 953-958.
3. Chamberlin, E. M.; F, P. B.; Williams, D. E.; Conn, J., J. Org. Chem. 1962, 27,
2263-2264.
4. Schiller, R. L.; Lincoln, S. F.; Coates, J. H., J. Chem. Soc., Faraday Trans. 1
1987, 83, 3237-3248.
5. Binstead, R. A.; Jung, B.; Zuberbuhler, A. D. SPECFIT/32, v3.0.39(b); Spectrum
Software Associates: Marlborough, MA, USA, 2007.
6. Gans, P.; Sabatini, A.; Vacca, A., Talanta 1996, 43, 1739-1753.
7. Gans, P.; Sabatini, A.; Vacca, A., Ann. Chim. (Rome) 1999, 89, 45-49.
8. Frassineti, C.; Ghelli, S.; Gans, P.; Sabatini, A.; Moruzzi, S.; Vacca, A., Anal.
Biochem. 1995, 231, 374-382.
9. Gans, P.; Sabatini, A.; Vacca, A. HypNMR, v3.1.5; Protonic Software: 2004.
10. CambridgeSoft, ChemBio3D Ultra, v11.0.1; Cambridge, 2007.
11. Stewart, J. J. P., J. Mol. Mod. 2007, 13, 1173-1213.
12. Stewart, J. J. P., MOPAC2009, v. 10.288w; Stewart Computational Chemistry:
2009.
13. Agrafidtis, D.; Rzepa, H., J. Chem. Res. Synop. 1988, 100-101.
14. Head, J.; Zerner, M., Chem. Phys. Lett. 1985, 122, 264-270.
15. Murakami, T.; Harata, K.; Morimoto, S., Tetrahedron Lett. 1987, 28, 321-324.
Huy Tien Ngo Chapter 5 - Experimental
208
16. Ueno, A.; Breslow, R., Tetrahedron Lett. 1982, 23, 3451-3454.
17. Breslow, R.; Czarnik, A. W., J. Am. Chem. Soc. 1983, 105, 1390-1391.
18. Murakami, T.; Harata, K.; Morimoto, S., Chem. Lett. 1988, 553-556.
19. Du, H.-T.; Du, H.-J.; Lu, M.; Sun, L.-L., Acta Cryst. 2007, E63, o4926.
20. Guo, K.; Chu, C. C., J. Polym. Sci. Pol. Chem. 2007, 45, 1595-1606.
21. Easton, C. J.; van Eyk, S. J.; Lincoln, S. F.; May, B. L.; Papageorgiou, J.;
Williams, M. L., Aust. J. Chem. 1997, 50, 9-12.
22. Reija, B.; Al-Soufi, W.; Novo, M.; Tato, J. V., J. Phys. Chem. B 2005, 109, 1364-
1370.
23. Gianneschi, L. P.; Kurucsev, T., J. Chem. Soc, Faraday Trans. 2 1974, 70, 1334-
1342.
24. Pham, D.-T.; Ngo, H. T.; Lincoln, S. F.; May, B. L.; Easton, C. J., Tetrahedron
2010, 66, 2895-2898.
25. Palin, R.; Grove, S. J. A.; Prosser, A. B.; Zhang, M.-Q., Tetrahedron Lett. 2001,
42, 8897-8899.
26. May, B. L.; Clements, P.; Tsanaktsidis, J.; Easton, C. J.; Lincoln, S. F., J. Chem.
Soc, Perkin Trans. 1 2000, 463-469.
27. Guo, X.; Abdala, A. A.; May, B. L.; Lincoln, S. F.; Khan, S. A.; Prud'homme, R.
K., Macromolecules 2005, 38, 3037-3040.
28. Guo, X.; Abdala, A. A.; May, B. L.; Lincoln, S. F.; Khan, S. A.; Prud'homme, R.
K., Polymer 2006, 47, 2976-2983.
209
APPENDIX A
PUBLICATIONS
Based on the research carried out during the period of PhD candidature
1. Ngo, H. T., Clements, P., Easton, C. J., Pham, D.-T., Lincoln, S. F., Supramolecular
Chemistry of Pyronines B and Y, β-Cyclodextrin and Linked β-Cyclodextrin Dimers,
Aust. J. Chem., 2010; 63, 687-692.
2. Pham, D.-T., Ngo, H. T., Lincoln, S. F., May, B. L., Easton, C. J., Synthesis of C6A-to-
C6A and C3A-to-C3A Diamide Linked γ-Cyclodextrin Dimers, Tetrahedron, 2010, 66,
2895-2898.