Novel catanionic vesicles from calixarene and single-chain surfactant

Article abstract of DOI:10.1039/c0cc01806f

The mixed system between p-sulfonatocalix[4]arene and tetradecyltrimethylammonium bromide forms unilamellar vesicles after sonication of the aqueous dispersion. Furthermore these vesicles can be stored, without use of lyoprotectants, by lyophilization and

Full text of DOI:10.1039/c0cc01806f

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Novel catanionic vesicles from calixarene and single-chain surfactantw
Vitor Francisco,a Nuno Basilio,a Luis Garcia-Rio,*a Jose R. Leis,a Eduardo F. Maquesb and
a
´ ´
Carlos Vazquez-Vazquez
Received 9th June 2010, Accepted 15th July 2010
DOI: 10.1039/c0cc01806f
The mixed system between p-sulfonatocalix[4]arene and tetra-
decyltrimethylammonium bromide forms unilamellar vesicles
after sonication of the aqueous dispersion. Furthermore these
vesicles can be stored, without use of lyoprotectants, by
lyophilization and then rehydration without change in size
or shape.
SC4 is a well known receptor for organic ammonium cations
in water5 and displays especially strong binding abilities for
these guests due to their p-rich cavities and to five negative
charges at pH 7. Though ammonium cations are typical
guests for SC4 hosts, reported studies on the complexation
of alkylammonium cations are scarce.6
In contrast to SC6HM, when 2 mM of SC4 is mixed with
TTABr, we observe the formation of a white dispersion at
TTABr concentrations between 0.1 mM and 50 mM, and a
precipitate zone near the charge neutrality. Below and above
this gap a clear solution is observed. Since this last behavior is
common in some catanionic systems, we carried out further
experiments to identify the aggregates formed in that region.
In the literature, it is known that the mixture between the
above calixarene and some biorelevant molecules, such as
aliphatic amines, polyamines and amino acid isomers, is
organized in bilayer-type structures in the solid state.7 In this
work we study the host–guest system in liquid state and the use
of conventional amphiphilic surfactants, which increase the
number of molecules that can form bilayer structures with the
calixarene.
The construction of well-defined structures in the nanometre
or micrometre length scale based on molecular self-assembly is
one of the most important challenges facing modern chemistry.
For instance, noncovalent interactions have been used to
obtain a wide-range of structured aggregates such as tubules,
fibers, micelles, vesicles, and disks through molecular self-
assembly of small organic compounds.1
Mixtures of anionic and cationic surfactants (catanionic
mixtures) offer an attractive approach for the construction
of complex self-assembled nanostructures. The formation of
spontaneous vesicles in mixtures of oppositely charged surfactants
was first demonstrated by Kaler2 and since then, intense
research has been devoted to the study of self-assembled
structures formed in catanionic surfactant systems.3 Globular
micelles, cylindrical micelles, long threadlike micelle, discs,
and large lamellar sheets have also been observed in some of
the aqueous cationic–anionic systems. The molecular
assemblies formed in these systems are mainly attributed to
a strong electrostatic association modulated by chain packing
interactions, which generally result in a reduced head-group
area promoting a dense packing of surfactant molecules in the
aggregate.
When a milky dispersion of 50 mM SC4/TTABr with molar
ratio 1 : 2.5 was examined under Nomarski light microscopy
between glass and cover slide, a high concentration of giant
vesicles (0.5–5 mm) was visible (Fig. 1), with the smaller ones in
fast Brownian motion. The presence of these very large vesicles
is the reason for the white opaque appearance of the dispersion
and was detected throughout the 0.1–50 mM range.
In the diluted region, after sonication of a dispersion
containing 2 mM of SC4 and 5 mM of TTABr, the sample
was studied by dynamic light scattering (DLS) and transmission
electron microscopy (TEM). As shown below our results are
We have recently demonstrated that when the anionic
surfactant is replaced with a non-aggregating and surface
inactive hexamethylated p-sulfonatocalix[6]arene (SC6HM)
the micellization of single chain trimethylammonium
amphiphiles is promoted at concentrations below the critical
micelle concentration (cmc) of neat surfactant.4 It was
suggested that the complexation of the cationic surfactant
with SC6HM yields a species with surface active properties
that induce aggregation at lower concentrations than that of
neat surfactant. Motivated by this observation, we decided to
study the mixed system formed by the most common
p-sulfonatocalix[4]arene (SC4) with tetradecyltrimethyl-
ammonium bromide (TTABr).
a
´
Departamento de Quımica Fısica, Facultad de Quımica, Universidad
de Santiago, 15782 Santiago, Spain. E-mail: luis.garcia@usc.es
´
´
b
´
Centro de Investigac¸a˜o em Quımica, Department of Chemistry,
Faculty of Science, University of Porto, Rua do Campo Alegre,
no. 687, P 4169-007 Porto, Portugal
w Electronic supplementary information (ESI) available: Materials,
Experimental details and Dynamic Light Scattering (DLS) data. See
DOI: 10.1039/c0cc01806f
Fig.
1 A light micrograph of a 50 mM white dispersion of
SC4/TTABr with a molar ratio 1 : 2.5, showing the presence of
polydisperse and very large vesicles. The smallest visible vesicle
appears in Brownian motion.
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compatible with the presence of unilamellar vesicles in the
solution. When the aqueous mixture of components in the
sample is examined by DLS two relaxation modes are
observed: a fast mode at short relaxation times, related to
the diffusion of large aggregates, and (within the low experimental
precision because of bad statistics for such slow fluctuations in
a limited sampling time) a slow mode at long relaxation times
which is independent of the scattering vector q, indicating that
it is a ‘‘viscoelastic’’ mode. The average diffusion coefficient of
the vesicles is obtained by fitting the angular dependence
relaxation times of the fast mode, D = (4.29 Æ 0.05) mm2 sÀ1
,
Scheme 1 Two possible inclusion modes of TTABr in SC4.
which corresponds to an average hydrodynamic radius of
ca. 57.2 Æ 0.7 nm (see the ESIw).
Table 1 Chemical shift changes (Dd, ppm) for the inclusion complex
formed between the surfactant TTABr and the p-sulfonatocalix[4]arene.
Negative values indicate up-field shift
The size distribution was also investigated by TEM, and the
experimental results are in agreement with the average size
obtained by DLS, showing that the vesicles are generally
smooth and spherical (see Fig. 2). We have also measured
the charge of these vesicles in Z-Sizer equipment and a
z-potential about À23 Æ 5 mV was obtained at 25 1C. This
value met our expectations since we worked with an excess of
negative charge due to the calixarene. The diameter obtained
by the Z-Sizer is 135 nm and it is in accordance with TEM and
DLS data.
ArH aromatic ArCH2Ar RCH3 RCH2a N+(CH3)3
p-SC4
TTABr
Complex 7.61
Dd 0.03
7.58
—
4.01
—
4.04
0.03
—
—
3.41
2.15
—
3.17
1.31
À1.86
0.87
0.85
À0.02 À1.26
ESIw). No change was observed within 4–5 days from
preparation; however, after 7 days, a new relaxation mode is
observed at longer times than the fast diffusive mode. The
relative amplitude of the new mode increases, while that of the
corresponding fast mode decreases; this feature is an
indication that the vesicles are either coalescing and growing
in size or flocculating.
To obtain more information on the structure of the
aggregates NMR spectra of neat SC4, neat TTABr and mixed
vesicles were performed. The assigned chemical shifts are listed
in Table 1. The terminal protons of the surfactant alkyl chain
are unaffected in the complex. However, in contrast, the
N(CH3)3 and the protons bonded to the alpha carbon (Ca)
show large changes in chemical shifts, compatible with an
inclusion complex where the surfactant polar head is located in
the aromatic cavity of the calixarene (Scheme 1). In Scheme 1
the calixarene is represented in the simplified shape, ‘‘cone’’
This behavior is quite usual since often the high curvature
vesicles are metastable aggregates and consequently, the size
distribution evolves with time to larger structures (e.g. lamellar
sheets). The initial aqueous dispersions may even show phase
separation. In addition, chemical or biological degradation
may also develop. Due to this issue, dispersions that contain
vesicles must be freshly prepared just prior to use. Since this
process is often poorly defined and difficult to control,8 this
procedure presents some disadvantages (i.e., when preparing
liposome/DNA complexes).
1
conformation, but the H NMR spectrum indicates that the
calixarene is exchanging rapidly in the NMR time-scale
between several possible conformations, since the ArCH2Ar
methylene protons give one singlet.
In order to study the stability of the vesicles, the evolution
of the relaxation time spectra was studied by DLS (see the
To circumvent colloidal instability and/or avoid degradation
and to allow for long-term storage of vesicles, water may be
removed through the most common and frequent method to
dehydrate, that is the freeze-drying technique.9 The complete
process to stabilize and store our vesicles can be summarized
in the following four points: (A) p-Sulfonatocalix[4]arene
(SC4) and tetradecyltrimethyl-ammonium bromide (TTABr)
are dissolved in water yielding a whitish dispersion. (B) The
aqueous cloudy dispersion is sonicated for approximately
30 min, and a homogenous clear solution is obtained.
(C) The solution is then frozen with liquid nitrogen, causing
ice crystals to nucleate and grow. Sublimation of ice yields the
freeze-dried powder. (D) The dried power is rehydrated again
with water (Fig. 2).
After sample lyophilization (step C), a white powder is
obtained that completely redisperses in water forming again
the vesicles, without any need of sonication. Usually simple
hydration of the dried vesicles powder does not completely
redisperse them in water, but produces a mixture of suspended
vesicles and larger aggregates and therefore requires the
sample to be sonicated again.10
Fig. 2 Schematic representation for vesicle storage method and
rehydration. Steps 1, 2 and 3 correspond to sonication, lyophilization
and rehydration, respectively. The TEM images (negatively stained
2% phosphotungstic acid, pH = 7) are before step 4 and after the
lyophilization (step 5).
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Consequently we can deduce that the structure of vesicles
formed when the solution is sonicated for the first time
maintains their organization when water escapes and later
enters the structure again. Also we can conclude that this
process does not significantly change the size of the vesicles as
one might expect. Usually the conventional vesicle structure is
lost during the freeze-drying process, and as a result the use of
carbohydrates is introduced.11 The sugar coating on the
surface of the vesicles results in a low molecular mobility,
which minimizes damage caused by the fusion process or
crystal formation after drying.10 To confirm that these vesicles
do not need any cryoprotectant we have performed a new set
of DLS measurements after rehydration (see the ESIw).
From the linear fit, the average diffusion coefficient obtained
is D = (3.44 Æ 0.06) mm2 sÀ1, which corresponds to an average
hydrodynamic radius of 71 Æ 1 nm. This value confirms that
when these vesicles are lyophilized and hydrated again they do
not yield the thermodynamically preferred lamellar phase
domains. With respect to the process of water leaving and
entering the mixed amphiphilic film, the effect cannot be
considered very remarkable since the water permeability of
usual lipids is very high, for instance, almost 10 orders of
magnitude larger than that of sodium ion.12 We can derive
from this fact that the inclusion complex between p-SC4 and
TTABr does not significantly change the transport of water
across the vesicle bilayer.
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This work shows a new type of catanionic vesicle, of which
the principal feature is its potential or ability to be stored and
rehydrated on demand without any significant change in size.
Further investigations need to be done related to other
water-soluble calixarene and surfactants, as well as in the
combined properties of calixarenes as macrocyclic hosts and
self-organizing systems able to form vesicles. The lack of
toxicity and immune response of calixarene derivates enable
new applications of these macrocycles in biomedical and
pharmaceutical sciences.13
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Financial support from Ministerio de Ciencia y Tecnologıa
´
of Spain (Projects CTQ2008-04420/BQU and MAT2008-
06503/NAN) and Xunta de Galicia (Projects PGIDIT07-
PXIB209041PR and INCITE08PXIB209049PR). V.F. and
N.B. acknowledge FCT (Portugal) for PhD Grants SFRH/
BD/43836/2008 and SFRH/BD/29218/2006
Notes and references
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This journal is The Royal Society of Chemistry 2010
Chem. Commun., 2010, 46, 6551–6553 | 6553