Thermotropic and lyotropic properties of long chain alkyl glycopyranosides. Part II. Disaccharide headgroups

Article abstract of DOI:10.1016/S0009-3084(00)00151-1

We have investigated the thermotropic and lyotropic properties of some long chain alkyl glycosides with disaccharide headgroups. The thermotropism was measured with polarising microscopy and additionally the lyotropism with the contact preparation method, Fourier-transform Infrared (FTIR) spectroscopy, X-ray diffraction and small angle neutron scattering. A broad thermotropic as well as lyotropic polymorphism was found. The compounds displayed thermotropic S(A) (lamellar) and cubic phases, and the investigation of the lyotropic phase behaviour led to the observation of inverted bicontinuous cubic V(II) phases, lamellar L(α) phases, normal bicontinuous cubic V(I) phases, normal columnar H(I) phases, normal discontinuous cubic I(I) phases and lyotropic cholesteric phases. The phases are discussed with respect to the chemical structures that have been varied systematically to derive structure-property relationships. (C) 2000 Elsevier Science Ireland Ltd.

Full text of DOI:10.1016/S0009-3084(00)00151-1

Chemistry and Physics of Lipids
106 (2000) 157–179
www.elsevier.com/locate/chemphyslip
Thermotropic and lyotropic properties of long chain alkyl
glycopyranosides. Part II. Disaccharide headgroups
H.M. von Minden a, K. Brandenburg b, U. Seydel b, M.H.J. Koch c,
V. Garamus d, R. Willumeit d, V. Vill a,*
a Institut fu¨r Organische Chemie, Uni6ersita¨t Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg, Germany
b Research Center Borstel, Di6ision of Biophysics, Parkallee 10, D-23845 Borstel, Germany
c European Molecular Biology Laboratory c/o DESY, Notkestr. 85, D-22603 Hamburg, Germany
d GKSS Research Centre, Max Planck Straße, D-21502 Geesthacht, Germany
Received 13 January 2000; received in revised form 4 April 2000; accepted 6 April 2000
Abstract
We have investigated the thermotropic and lyotropic properties of some long chain alkyl glycosides with
disaccharide headgroups. The thermotropism was measured with polarising microscopy and additionally the
lyotropism with the contact preparation method, Fourier-transform Infrared (FTIR) spectroscopy, X-ray diffraction
and small angle neutron scattering. A broad thermotropic as well as lyotropic polymorphism was found. The
compounds displayed thermotropic SA (lamellar) and cubic phases, and the investigation of the lyotropic phase
behaviour led to the observation of inverted bicontinuous cubic VII phases, lamellar La phases, normal bicontinuous
cubic VI phases, normal columnar HI phases, normal discontinuous cubic II phases and lyotropic cholesteric phases.
The phases are discussed with respect to the chemical structures that have been varied systematically to derive
structure–property relationships. © 2000 Elsevier Science Ireland Ltd. All rights reserved.
Keywords: Alkyl glycopyranosides; Thermotropic liquid crystals; Lyotropic liquid crystals; Cholesteric phases; Cubic phases;
Glycolipids
Helferich (1911) was the first indication of ther-
motropic liquid crystalline properties in am-
1. Introduction
phiphilic carbohydrates.
These amphotropic molecules form both ther-
motropic liquid crystalline phases in their pure
state upon heating and lyotropic liquid crystalline
phases upon addition of a solvent. The first obser-
vation of lyotropic behaviour is also related to
studies on alkylated carbohydrates while
analysing the extracts from tuberculosis bacteria.
The observation of a double melting of certain
long chain alkyl glucopyranosides, e.g. hexadecyl-
b-D-glucopyranosoide (Fig. 1) by Fischer and
* Corresponding author. Tel.: +49-40-428384269; fax: +
49-40-428384325.
E-mail address: vill@liqcryst.chemie.uni-hamburg.de (V.
Vill).
0009-3084/00/$ - see front matter © 2000 Elsevier Science Ireland Ltd. All rights reserved.
PII: S0009-3084(00)00151-1

158
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
Koch (1884) observed unusual optical textures of
their aqueous dispersions.
solvents, the most common of which is water,
because of its biological relevance. Whereas small
amounts of water will not change the phase type
and transition temperatures, larger amounts will
introduce new phases such as cubic phases and
columnar phases (HI/II). The discontinuous cubic
phases are based upon various packings of spheri-
cal or slightly anisotropic micelles, whereas the
bicontinuous cubic phases with interwoven fluid
porous structures are based upon underlying infi-
nite periodic minimal surfaces (Fairhurst et al.,
1998).
This phase in particular is of great biological
interest (Siegel, 1986a,b,c; Siegel et al., 1989; El-
lens et al., 1989; Lindblom and Rilfors, 1989). The
lamellar phase with its multibilayer structure has
long been well known to the biologists, but there
is much evidence today that the bicontinuous
cubic phase may play an important role in pro-
cesses such as membrane fusion.
Whereas the first observation of liquid crys-
talline properties of alkylglycosides was related to
long chain compounds, subsequent interest fo-
cused only on their shorter chain counterparts
(Jeffrey, 1984; Jeffrey and Wingert, 1992; Prade et
al., 1995) because of their interesting properties as
surfactants (Bo¨cker and Thiem, 1989; Vill et al.,
1989; Boullanger, 1998). These compounds dis-
solve in water, whereas the long chain glycosides
do only swell.
The driving force for the mesophase formation
in the case of amphiphilic molecules is a micro
phase separation leading to an aggregate structure
with separated regions for the lipophilic and hy-
drophilic molecular moieties. This structure en-
ables the maintenance of the van der Waals
interaction in the hydrophobic region and of the
hydrogen bonding in the hydrophilic region, each
stabilising the formed mesophase.
The principal phase behaviour of these com-
pounds is shown in Fig. 2 (Prade et al., 1995;
Blunk et al., 1998). A typical amphiphile is repre-
sented by a vertical line and usually exhibits only
one mesophase, but in the case of an unusual
geometry of the hydrophilic headgroup polymor-
phism can occur. In this case, cubic phases may
also be observed (Fischer et al., 1994), while
simple amphiphiles such as the one shown in Fig.
1 normally exhibit thermotropically only an SA
phase (Jeffrey and Wingert, 1992).
Deviation from the vertical line mentioned
above may also be seen upon the addition of
We synthesised long-chain alkyl glycopyra-
nosides (in particular stearyl glycosides) which
can be incorporated in biomembranes, and
change the sugar headgroup systematically to elu-
cidate the structure–property relationships gov-
erning the occurrence of the polymorphism
mentioned above. In the case of a disaccharide
headgroup, we observed a broad range of phases.
Fig. 1. Hexadecyl-b-D-glucopyranoside.
2. Materials and methods
2.1. Materials
General reaction conditions were as follows.
The alkyl glycosides had been synthesised as
shown in Fig. 3 for the cellobiosides using the
method described by Vill et al. (1989), however
Fig. 2. Principal phase behaviour of amphiphilic compounds.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
159
2.2.1. Sample preparation for lyotropic phase
beha6iour
The glycolipid samples were prepared as
aqueous dispersions at 90% buffer content using
20 mM Hepes, in some cases at lower water
content (50%) also. For this, the lipids were di-
rectly suspended in the buffer, and the suspen-
sions were temperature-cycled several times
between 5 and 70°C and then stored for at least
12 h at 4°C before measurement.
2.2.2. FTIR spectroscopy
Infrared spectroscopic measurements were per-
formed on FTIR spectrometers ‘5-DX’ (Nicolet
Instruments, Madison, WI) and IFS-55 (Bruker,
Karlsruhe, Germany). The lipid samples were
placed in a CaF2 cuvette separated by a 12.5-mm
thick teflon spacer. Temperature scans were per-
formed automatically between 10 and 70°C with a
heating rate of 3°C per 5 min. Every 3°C, 50
interferograms were accumulated, apodised,
Fourier-transformed, and converted to ab-
sorbance spectra. The gel to liquid crystalline
phase transition LblLa (SB to SA in lamellar
phases) was determined by evaluating the sym-
metric stretching vibration of the methylene
groups ns(CH2) with standard procedures. The
location of ns(CH2)-around 2850 cm−1 in the gel
phase and 2853 cm−1 in the fluid phase — is a
sensitive indicator of acyl chain order (Casal and
Mantsch, 1984).
Fig. 3. Synthesis of alkyl glycopyranosides.
boron trifluoride diethyl etherate was used instead
of tin tetrachloride as Lewis acid for the synthesis
of the b-anomers. Furthermore, no ion exchange
chromatography was necessary in our case to
separate the anomers, since a silica gel chro-
matography [light petroleum (b.p. 60–70°C)–
ethyl acetate 4:1] was already successful, while in
the case of the homologous dodecyl glycosides
prepurification by silica gel chromatography and
a subsequent ion exchange chromatography after
deprotection on Dowex 1×2 has been used (Ro-
sevar et al., 1980; Bo¨cker and Thiem, 1989). For
further information concerning the synthesis and
characterisation, please contact the author of
correspondence.
2.2.3. X-ray diffraction
X-ray diffraction measurements were per-
formed at the European Molecular Biology Labo-
ratory (EMBL) outstation at the Hamburg
Synchrotron Radiation Facility HASYLAB using
the double-focusing monochromator-mirror cam-
era X33. Diffraction patterns in the range of the
scattering vector 0.07BsB1 nm−1 (s=2 sin
q/u, 2q the scattering angle, and u the wave-
length=0.15 nm) were recorded at 40°C with
exposure times of 2 or 3 min using a linear
detector with delay line readout (Gabriel and
Dauvergne, 1982). The s-axis calibration was
done using tripalmitin as standard (periodicity
4.06 nm at 25°C). Further details of the data
acquisition and evaluation system can be found
2.2. Methods
An Olympus BH optical polarising microscope
equipped with a Mettler FP 82 hot stage and a
Mettler FP 80 central processor was used to iden-
tify thermal transitions and characterise an-
isotropic textures.

160
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
elsewhere (Boulin et al., 1986). In the diffraction
patterns presented here, the logarithm of the dif-
fraction intensities log I(s) is plotted against the
scattering vectors. The evaluation of the X-ray
spectra was made according to procedures de-
scribed by Luzzati et al. (1986) and in previous
papers (see Brandenburg et al., 1990, 1998) which
allow the assignment of the spacing ratios of the
main scattering maxima to defined three-dimen-
sional structures. Here lamellar and cubic struc-
tures are of particular relevance, which can be
characterised by the following features.
procedures (Stuhrmann, 1989). The two-dimen-
sional isotropic scattering spectra were az-
imuthally averaged, converted to an absolute
scale, and corrected for detector efficiency by
dividing by the incoherent scattering spectra of
pure water (Wignall and Bates, 1986) which was
measured with a 1-mm-path-length quartz cell.
2.2.5. SANS analysis
2.2.5.1. Model-independent approach. The pair dis-
tribution function, p(r), of scattering length den-
sity of studied micelles is in principle obtained by
Fourier transformation of the measured difference
cross-section of neutron scattering. The overall
shape and dimensions of the micelles can in many
cases be estimated from the pair distribution func-
tion (Glatter and Kratky, 1982.). As the scattering
is only measured in a finite q range, a traditional
Fourier transformation cannot be performed. In-
stead, the method of indirect Fourier transforma-
tion is used (Glatter, 1977; Pedersen, 1997).
Having determined p(r) we are able to calculate
integral parameters of the micelle (Glatter, 1992)
such as the radius of gyration Rg.
1. Lamellar (smectic phases)
The reflections are grouped in equidistant
ratios, i.e., at 1, 1/2, 1/3, 1/4 etc. of the
lamellar repeat distance dl.
2. Cubic
These are non-lamellar three-dimensional
structures. Their various space groups differ
in the ratio of their spacings. The relation
between reciprocal spacing shkl=1/dhkl and
lattice constant a is shkl=(h2+k2+l2)1/2/a
(hkl=Miller indices of the respective set of
plane).
In addition, X-ray wide-angle diffraction was
performed in selected cases, in which the determi-
nation of the short-range order is possible. In the
case of a hexagonal array of the lipid chains such
as found for many phospholipids, the distance
between neighbouring acyl chains, the lattice con-
stant a, can be calculated from the main wide-an-
gle reflection at a spacing ratio d by a=2/ꢀ3d.
2.2.5.2. Modelling. The difference cross section of
neutron scattering of the population of non-inter-
active particles (Glatter and Kratky, 1982) can be
written as
dS (q)
dV
dS (0)
=
,
dVF2(q)+Binc
2.2.4. Small angle neutron scattering
A set of small-angle neutron scattering experi-
ments was performed on the SANS1 instrument
at the FRG1 research reactor of GKSS,
Geesthacht, Germany (Stuhrmann et al., 1995).
The range of scattering vectors q from 0.0008 to
0.025 nm−1 was covered by three combinations of
neutron wavelength (0.85 nm) and sample-to-de-
tector distances (from 0.7 to 7 m). The wavelength
resolution was 10% (full-width-at-half-maximum
value).
where dS (0)/dV is the scattering at ‘zero’ angle
and is related to concentration of particles, vol-
ume and difference of scattering length densities
between particle and solvent, Binc is the residual
incoherent background. The form factor F(q) ex-
presses the scattering cross section of particle. In
the case of particle of ellipsoidal or cylindrical
shape (Pedersen, 1997), it is written as
y/2
&
F2(q)=
[ f(q,i)]2 sin idi
The samples were kept in quartz cells with a
path length of 1.5 mm at 23°C. The raw spectra
were corrected for backgrounds from the solvent,
sample cell, and other sources by conventional
0
In the case of a homogeneous cylinder f(q, i) is
written as

162
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
Fig. 4. (Continued)
sin (qL/2 cos i)2J1(qR sin i)
The models mentioned above were applied to fit
experimental data.
f(q,i)=
,
(qL/2 cos i)(qR sin i)
where L is the length of the cylinder, R is the
radius of the cylinder, J1 is the first-order Bessel
function, and i is the angle between the q vector
and the axis of the cylinder.
In the case of an ellipsoid of rotation with semi
axis R, R, mR f(q, i) is written as
3. Results and discussion
3.1. Thermotropic phase beha6iour
Fig. 4 depicts a systematic structure variation
3[ sin (x)−x cos (x)]
of octadecyl-b- -glucopyranoside 2. While the
D
f(q,i)=
(x)3
first part of this paper (Vill et al., 1999) focussed
on long chain alkyl glycopyranosides with
monosaccharide headgroups, we present here re-
where x=qR[ sin2 i+m2 cos2 i]1/2
.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
161
Fig. 4. Thermotropic behaviour of the synthesised amphiphilic liquid crystals; (a) the compound formed a glass and, thus, no
melting point could be determined; (b) the synthesis of this compound and the rough transition temperatures without a
characterisation of the phase type were reported first bei Hori (1958); (c) an additional (probably columnar) metastable mesophase
was formed on cooling.

164
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
Fig. 5. (Continued)
sults obtained by replacing the monosaccharide
moiety of compound 2 by disaccharides.
seems that the effect of structural changes in the
aliphatic chain on the clearing temperature are
stronger in the case of monosaccharide head-
groups than in the case of disaccharide head-
groups. An interesting observation, which further
supports this hypothesis, is that of the effect on
the clearing temperature, if the oxygen is replaced
by sulphur as linking atom between the hy-
drophilic and hydrophobic moieties of a malto-
If the glucose headgroup of compound 2 is
replaced by the a 1“4-linked disaccharide unit of
maltobiose (compound 4), the clearing tempera-
ture increases dramatically by 127 K due to in-
creased hydrogen bonding in the headgroup
region attributable to the increased number of
hydroxyl groups. Furthermore, a better ratio be-
tween the polar and unpolar moieties is obtained
by exchanging the glucose headgroup of 2 for a
disaccharide unit. The optimal hydrophilic/hydro-
phobic balance and, therefore, the highest clearing
temperature is reached in the case of monosaccha-
ride headgroups with a chain of 12–14 carbon
atoms (Prade et al., 1995). However, the optimal
balance between the hydrophilic and hydrophobic
moieties is reached in case of disaccharide head-
groups at longer chain length.
The introduction of a unsaturation in com-
pound 6 again lowered the clearing temperature,
but with only 7 K less than in the case of the
monosaccharide derivatives described in part I of
this paper. The reason for the lower clearing
temperatures of the unsaturated derivatives in
general can be seen in the disturbance introduced
into the alkyl chain region by the double bond,
circumventing a close packing of the alkyl chains
and, therefore, lowering the van der Waals inter-
actions. The stronger effect of the unsaturation on
the monosaccharide derivatives with an already
imbalanced ratio between the polar and unpolar
molecular moieties might be explained by the
broadening of the alkyl chains introduced through
the unsaturation, increasing the hydrophobic vol-
ume and hence the imbalance even further. How-
ever, there is still a more general explanation: it
bioside. In the case of a-
Doren et al., 1989) [unfortunately insufficient data
are available about the b- -thioglucosides] and
D-thioglucosides (Van
D
also in the case of many other thio compounds
with a monosaccharide headgroup-e.g. the 1-thio-
glycerides (Van Doren et al., 1990), 1-alkylthio-1-
deoxy-
galacturonic acid (Vogel et al., 1992) and dialkyl
thioacetals of -glucose (Dahlhoff, 1987; Eckert
D-glucitols (Dahlhoff, 1990), thioesters of
D
et al., 1987; Van Doren et al., 1988; Tietze et al.,
1994), the thio derivatives always exhibit distinctly
higher clearing temperatures than the oxygen-
linked derivatives. Nevertheless, the dodecyl thio
maltobioside 8 exhibits nearly the same clearing
temperature as the oxygen-linked maltobioside
(compound 35, Fig. 5) and proves, therefore, our
hypothesis. For other reasons, we also synthesised
the tridecyl thio maltobioside 10 with a clearing
temperature of 260.0°C. Unfortunately, no litera-
ture data are available on the clearing tempera-
ture of the analogous oxygen-linked compound,
but we calculated a value of 254.5°C using
LiqCryst data base (Vill, 1999). This value seems
reasonable, since the maltobiosides do not show
an odd–even effect, and the calculated tempera-
ture should, thus, lie in the middle of the clearing
temperatures of the dodecyl and tetradecyl malto-
bioside (Fig. 5, compounds 35 and 36). This tem-

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
163
Fig. 5. Literature data for the transition temperatures of some amphiphilic liquid crystals for comparison; (a) in the literature no
exact melting point was given, since the compounds formed non-crystalline solids after lyophilisation.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
165
perature is again only slightly lower than the
clearing temperature of the thio derivative 12.
In the next step, we changed the type of the
interglycosidic linkage (a vs. b), the position of
linkage (1“4 vs. 1“6), and the configuration of
the second sugar residue (glucopyranosyl vs.
galactopyranosyl) systematically. The clearing
temperatures of these compounds varied over a
range of 33 K owing to the various kinds of
configurations, and in several cases cubic phases
were found.
group. The difference in stability between the two
anomers is in case of the cellobiosides, e.g.
cel-b-OC18 (14) vs. cel-a-OC18 (12) 20.5 K
cel-b-OC12 (39) vs. cel-a-OC12 (38) 35.4 K
and similar results are found for the
maltobiosides,
mal-b-OC14 (36) vs. mal-a-OC14 (41) 33 K
mal-b-OC12 (35) vs. mal-a-OC12 (40) 40 K.
3.1.2. Influence of the type of linkage between the
two sugar residues (h 6s. i)
3.1.1. Influence of the type of linkage (h 6s. i) of
the aliphatic chain to the disaccharide headgroup
In all these cases, the compounds with the
aliphatic chain linked by a b glycosidic bond
(compounds 14, 18, 22, 28, 30 and 34) to the
disaccharide moiety displayed a higher clearing
temperature (and also the higher m.p.) and,
hence, the more stable smectic A phase than the
corresponding a anomer (12, 16, 20, 26, 37 and
32). This difference in stability was in the case of
compounds with disaccharide headgroups with a
interglycosidic 1“6 linkage with about 10 K
(compounds 22, 28, 30 and 34 vs. 20, 26, 37 and
32) smaller than in the case of the compounds
with the two glycosyl units of the disaccharide
headgroup being linked by a 1“4-linkage (about
20 K, compounds 14 and 18 vs. 12 and 16). The
reason for the greater stability of the compounds
If the a 1“4-linked second glucopyranosyl
residue of the maltoside 4 is replaced by a b
1“4-linked glucopyranosyl residue (cellobiosides
14), the clearing temperature increases by 9.5 K
due to the stiffer and more rod-like structure of
the cellobioside headgroup. This difference in sta-
bility also increases again with decreasing chain
length as can be seen easily from the data in
Fig. 5.
Whereas the maltobiosides displayed only
smectic phases, the a cellobioside 12 displays a
monotropic cubic phase (see also Table 1), which
could not be found in the case of the b cel-
lobioside 14, since on cooling of the SA phase
crystallisation occurred already some degrees be-
low the melting point. These results will be dis-
cussed together with the lactosides in the
following section.
with
a b-anomerically-linked aliphatic chain
might be seen in their somewhat more rod-like
shape. This effect is not so strong in the case of
compounds with a 1“6-linked disaccharide unit,
since the sugar moiety is angular and causes the
molecule already to be banana-shaped. This devi-
ation from a rod-like structure is much stronger
than the one introduced by an a linkage of the
aliphatic chain to the sugar moiety, and a further
deviation from a rod like structure due to the
latter, therefore, decreases the stability of the SA
phase only slightly.
3.1.3. Influence of the configuration of the second
sugar residue (glucopyranosyl 6s.
galactopyranosyl)
If the second, b 1“4-linked glucopyranosyl
residue of the cellobiosides 12 and 14 is replaced
by a b 1“4-linked galactopyranosyl residue, the
stability of the SA is more or less not effected; the
clearing temperature of the a lactoside 16 is 2.8 K
lower than the clearing temperature of the corre-
sponding a cellobioside while the clearing temper-
ature of the b lactoside 18 is 0.5 K higher than in
the case of the b cellobioside. The configuration at
C-4 of the second sugar moiety seems to have,
therefore, more or less no influence on the stabil-
ity of the SA phase in the case of 1“4-linked
While this difference in stability between the
two anomers is more or less chain-length indepen-
dent in case of a 1“6-linked headgroup (meli-
biosides, 22 and 28 vs. 20 and 26), it is strongly
dependent on the chain length in case of com-
pounds with a 1“4-linked disaccharide head-

166
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
disaccharide headgroups, but this is not true in
the case of 1“6-linked disaccharide headgroups
(see below).
The exact structure of the headgroup seems to
have an influence on the occurrence of this
monotropic cubic phase (even if it cannot be
understood yet), since the a cellobioside 12, the a
and b lactobiosides 16 and 18, and the b gento-
bioside 24 exhibited a monotropic cubic phase
while it was not found in the case of the b
maltobiosides 4 and 6 and the a and b meli-
biosides 20 and 22. At present, only two very
general structural requirements can be derived,
“ a disaccharide headgroup seems to be neces-
sary, since the similar stearyl gluco- and galac-
topyranosides (see first part of this paper) did
not show this unusual mesogenic behaviour
“ a chain of sufficient length is also necessary,
since e.g. the homologous dodecyl cellobiosides
or tetradecyl lactosides (Vill et al., 1989) only
displayed a SA phase.
Rod-like amphiphilic molecules are known to
display a SA phase, the stability of which increases
the more balanced the ratio between the polar and
apolar moieties becomes, and decreases after-
wards again. If this imbalance between the hy-
drophilic and hydrophobic moieties is increased
sufficiently, a cubic phase might be observed in-
stead of the SA phase usually found. These find-
ings are similar to the results obtained earlier with
some glucamine derivatives (sodium salts of N-
The a lactoside 16 as well as the b lactoside 18
displayed, as did the
a cellobioside 12, a
monotropic cubic phase. The introduction of the
galactopyranosyl residue improved in case of the
b lactoside compared to the b cellobioside the
supercoolability and inhibited the crystallisation,
which prevented the occurrence of a monotropic
cubic phase of the b cellobioside 14. So far, only
a few sugar derivatives are known to show a
thermotropic cubic phase (e.g. Praefcke et al.,
1990; Fischer et al., 1994; Beginn et al., 1997;
Borisch et al., 1997Borisch and Diele, 1997) and
one of them is the dodecyl-a- -gentobioside 37
D
(Fischer et al., 1994), some homologous com-
pounds of which have been synthesised in this
work and will be discussed below. The occurrence
of cubic phases might be attributable in these
cases to their banana-shaped molecular structures,
whereas the occurrence of thermotropic cubic
phases in the case of cello- and lactobiosides is
rather unexpected and difficult to explain, since
compounds of this type are so far only known to
form SA phases according to our knowledge (Jef-
frey and Wingert, 1992; Prade et al., 1995). Be-
cause of their rod-like molecular shape they are
indeed something like the prototype of SA phase
forming-amphiphilic glycosides.
alkyl-N-carboxymethyl-D-glucamine, Vill et al.,
1992), but it is still impossible to explain the
Table 1
Gel to liquid crystalline phase transition Tc, type of supramolecular aggregate structures (L lamellar, Q cubic, and H hexagonal),
and position of wide-angle X-ray diffraction for various ‘glu–glu’ and ‘glu–gal’ glycolipids
Glycolipid
Tc (°C)
Aggregate structures
Wide-angle reflection (nm)
Mel-b-OC12 26
Mel-b-OC12 28
Mel-b-OC18 20
Mel-b-OC18 22
Mal-b-SC12 8
Mal-b-SC13 10
Mal-b-OC18 4
Mal-b-OC18:1 6
Cel-b-OC18 12
Cel-b-OC18 14
Lac-b-OC18 16
Lac-b-OC18 18
Gen-b-OC18 24
L
L
No reflection
No reflection
0.451, unsharp
n.m.
No reflection
0.441 and 0.256
0.424 (5°C), 0.420 (20–40°C)
No reflection
0.466 and 0.390
n.m.
B0
38
67.5
B0
6
40
B0
36 (Tpre) 56
70
53(Tpre) 70
78
54
Q(5–40°C) L/unknown (60–80°C)
L
L (5–4°C) L/L’ interdigitated (60–80°C)
L
HI\L\Q
L
Q (5–60°C), L(80°C)
L
L
L
0.442
0.453, unsharp
n.m.
L(5–60°C), HII (80°C)

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
167
The most obvious difference between the gly-
cosides with a 1“6- and a 1“4-linked disaccha-
ride headgroup is the more or less abundant
occurrence of cubic phases in the former case.
Except for the dodecyl-a-isomaltoside 32, all of
the dodecyl glycosides (26, 28, 30, 34 and 37) with
a 1“6--linked disaccharide headgroup exhibit a
cubic mesophase. Since the a melibioside 26
showed a cubic mesophase while the a isomal-
toside 32 did not, this difference in the mesogenic
behaviour must be attributed to the changed
configuration at C-4 of the second sugar residue
due to the exchange of a galactopyranosyl for a
glucopyranosyl residue.
Fig. 6. Types of amphiphiles.
influence mentioned above of the sugar
headgroup.
3.1.4. Influence of the position of the linkage
between the two sugar moieties in the
disaccharide headgroup (1“6 6s. 1“4)
The occurrence of a cubic mesophase might be
explained in these cases by the broad structure of
the 1“6-linked disaccharide headgroups, giving
the molecules a banana-shaped structure (this
term has been used before by Takezoe (Niori et
al., 1996) to describe the molecular shape of a
completely different class of symmetric non-car-
bohydrate liquid crystals with different properties
than the compounds discussed here).
While the 1“4-linked disaccharides are always
more or less linear, the 1“6-linked disaccharides
are generally angular, yielding a banana-shaped
structure of the glycosides made thereof, which
influences their mesogenic behaviour strongly.
The stability of the SA phase is reduced due to the
deviation from a rod-like shape as can be seen
from a comparison with the transition tempera-
tures of the analogous 1“4-linked derivatives.
The extent to which the stability of the SA phase
is reduced depends strongly on the length of the
alkyl chain e.g.
Compounds of the general structure A (Fig. 6)
have so far only been reported to display smectic
phases (Jeffrey and Wingert, 1992; Prade et al.,
1995), and compounds of type B show columnar
phases with the interfacial area being curved
around the minor component (Mannock et al.,
1987, 1990 Jeffrey and Wingert, 1992; Prade et al.,
1995). In the case of compounds of type C, the
headgroup prefers a columnar structure while the
aliphatic tail favours a smectic structure, but each
is destabilised by the other moiety. The discussed
glycosides with a 1“6-linked disaccharide head-
group belong to this last group, which as a result
of frustration forms cubic mesophases as a com-
promise. The cubic to SA transition may be at-
tributed to the increasing thermal motion in the
alkyl chains, leading to a greater effective volume
of the hydrophobic chain. This causes the
molecule to become rod-like at elevated tempera-
tures and, hence, a cubic to SA transition is found.
If the dodecyl chain is extended, as in the case
of the stearyl melibiosides (20, 22), only an SA
phase is observed, whereas the stearyl gentio-
bioside 24 displays a monotropic cubic phase.
This difference in behaviour might be explained
gen-b-OC18 (24) vs. cel-b-OC18 (14): 15.5 K
mel-b-OC18 (22) vs. mal-b-OC18 (4): 12.0 K
(because an exchange of the second glucopyra-
nosyl residue in the cellobioside for a galactopyra-
nosyl residue did not effect the clearing
temperatures, it seems possible to compare the
melibiosides with the maltobiosides instead of
comparing with the proper compounds with a
disaccharide headgroup of a galactopyranosyl
residue linked by an a 1“4 bond to a glucopyra-
nosyl residue)
gen-b-OC12 (30) vs. cel-b-OC12 (39): 91.4 K
mel-b-OC12 (28) vs. mal-b-OC12 (35): 74.0 K
isomal-b-OC12 (34) vs. mal-b-OC12 (35):
91.0 K

168
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
“ There are two different structural features
which support the occurrence of thermotropic
cubic mesophases,
by the difference in size of the sugar headgroups.
With the melibiose headgroup, the stearyl gly-
cosides apparently have a rod-like shape at all
temperatures, whereas in the case of the gentio
bioside it becomes rod-like at elevated tempera-
tures only.
These two cubic phases (the one observed with
banana-shaped short chain molecules and the one
observed with long chain glycosides) probably
belong to two different groups as might be con-
cluded from the contact preparations with water
(see below), the former to the normal type of
bicontinuous cubic phases and the latter to the
inverted type. These terms are normally only used
to describe the structure of lyotropic cubic
mesophases, but it seems likely that they can be
extended to describe these structures. X-ray inves-
tigations and model simulations performed earlier
to investigate the structure of the cubic mesophase
ꢀ linear structures with an extremely imbal-
anced ratio between the hydrophilic and
hydrophobic
moieties
may
form
monotropic cubic mesophases.
ꢀ extremely bent molecular structures, as
they are found in glycosides with a 1“6-
linked disaccharide moieties, form cubic
mesophases in the case of an appropriate
length of the aliphatic chain.
3.2. Lyotropic phase beha6iour
3.2.1. Structural lyotropism
Above the critical micellar concentration
(CMC), glycolipids form supramolecular aggre-
gates in aqueous media. The type of aggregate
structure can be assumed to play a biologically
relevant role and is determined by the conforma-
tion (shape) of the contributing molecules, which
is determined by their primary chemical structure
and is influenced by ambient conditions such as
pH and concentration of mono- and divalent
cations (Israelachvili, 1991). Moreover, the molec-
ular shape of a given glycolipid molecule depends
on the state of order of the chains, which can
assume two main phase states, the gel (b-phase)
and the liquid crystalline (a-phase) states. Be-
tween these two-phase states, a reversible transi-
tion can take place at a given phase transition
temperature Tc. The value of Tc is dependent in
the first place on the number, length, and degree
of saturation of the acyl chains, the conformation
and the charge density and its distribution within
the headgroup region, and the nature and size of
the saccharide moiety. A closer characterisation
of the main types of supramolecular aggregate
structures (micellar, lamellar L, cubic Q and in-
verted hexagonal HII) occuring in glycolipid:water
systems is given by (Curatolo, 1987). More gen-
eral information about structural polymorphism
can be found elsewhere (Luzzati et al., 1986;
Seddon, 1990; Israelachvili, 1991; Seydel et al.,
1993). A comprehensive overview of the phases
and phase transitions (including the three-dimen-
sional aggregate structures L, Q, and HII) adopted
of dodecyl-a- -gentiobioside 37 (Fischer et al.,
D
1994) point towards a cubic phase of the space
group No. 230, Ia3d, made up by a network of
cylinders like in the lyotropic case.
At the end of this section, it seems useful to
summarise the results of the discussion of the
thermotropic properties.
“ The disaccharide derivatives show generally
higher melting and clearing points due to the
increased hydrogen bonding in the headgroup
region.
“ The influence of structural changes to the
aglycon is diminished in the case of disaccha-
ride derivatives compared with the monosac-
charide compounds.
“ The stability of the SA phase increases, the
more rod-like the sugar headgroup becomes,
generally favouring the b linkage of the sac-
charide moiety to the aliphatic chain as well
as the b linkage between the two glycosyl
residues. For the same reason, the stability of
the SA phase of compounds with a 1“6-
linked disaccharide moiety is decreased com-
pared with their counterparts with
a
1“4-linked disaccharide headgroup.
“ The difference mentioned above in stability
between the anomers decreases with increas-
ing length of the aliphatic chain.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
169
Fig. 7. Phase sequences observed in the contact preparation of the synthesised amphiphilic liquid crystals with water; (a) a cubic
glass was formed on cooling of the melted sample, which was used for the contact preparation; (b) the boundary of the HI phase
towards the water region was somewhat diffuse, thus, the existence of a lyotropic cholesteric phase could not be proved without
doubts.

170
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
by glycoglycerolipids is given by Koynova and
Caffrey (Koynova and Caffrey, 1994).
of this paper; Vill et al., 1999). Myelins are mobile
tubular structures, which are considered to consist
of cylindrical or helical arrangements of many
(300–5000) of the bilayers that constitute the
lamellar phase (Jeffrey and Wingert, 1992; Bouli-
gand, 1998).
The formation of lyotropic phases was investi-
gated by means of X-ray diffraction, small-angle
neutron diffraction and the contact preparation
technique (Van Doren and Wingert, 1994). If
water penetrates the sample, one or more ly-
otropic phases are formed, which can be identified
by polarising microscopy based on their charac-
teristic textures. Because a gradient of water con-
centration is generated, the whole range of phases
starting from the anhydrous bulk in the middle to
the completely hydrated sample at the outermost
region can be observed. Depending on the posi-
tion of a particular phase with respect to the other
phases, it is even possible to classify it tentatively
as belonging to the normal or inverted type in the
case of cubic or columnar phases; for the same
reason, it is possible to differentiate between a
bicontinuous and a discontinuous (micellar) cubic
phase. For example, a cubic phase, observed at a
higher water concentration than the lamellar
phase and followed by a columnar phase should
be of the normal bicontinuous type.
If the monosaccharide headgroup of 2 is re-
placed by the disaccharide maltobiose (compound
4), three phases are formed with increasing water
concentration, a lamellar La phase; a columnar HI
phase; and a further, only weakly anisotropic
phase. Replacing the stearyl chain by an oleyl
chain (6) leads to the appearance of an additional
broad bicontinuous cubic VI phase, and this phase
sequence, i.e. La, VI, HI, and a weakly anisotropic
phase, is also found in the case of the dodecyl-
and tridecyl thiomaltosides 8 and 10. The already
mentioned weakly anisotropic phase, which ap-
peared as a more or less thin band at the edge of
the HI phase towards the water region, was also
found with all other compounds except for the
dodecyl a and b melibiosides 26 and 28 (Fig. 7).
Since a micellar cubic II phase can be ruled out as
explanation because of its isotropic structure, a
lyotropic cholesteric phase might be supposed as
explanation. This phase is expected to appear at
higher water concentration than the columnar HI
phase, and to be anisotropic. Both of these crite-
The stearyl-b- -glucopyranoside 2 displayed in
D
a contact preparation with water only myelin
figures as did other long chain alkyl glycosides
with a monosaccharide headgroup (see first part
Fig. 7. (Continued)

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
171
Fig. 8. Distance distribution function p(r) calculated from the experimental SANS data. The experimental solutions were prepared
by dissolving the maltobiosides 8 and 10 in heavy water at a concentration of 90 mmol and 100 mmol, respectively.
ria are fulfilled by the phase that has been found.
Whereas quite a lot is known about thermotropic
cholesteric phases, their lyotropic counterparts are
still seldom observed and not well understood.
The mesophase of lyotropic cholesteric phases is
built up by anisotropic micelles.
kinds of micelles. It can easily be seen that the
experimental data match very well with the model
fits for cylindrical micelles, and with this informa-
tion the size of the micelles forming the lyotropic
cholesteric phase of compounds 8 and 10 was
calculated from the SANS data.
Compound 8
Radius of cylinder 2.0090.01 nm, Length
29.090.1 nm.
The size and shape of the micelles was deter-
mined in the case of the dodecyl- and tridecyl-b-
D
-thiomaltobioside 8 and 10 by small-angle
Compound 10
neutron scattering (SANS).
Radius of cylinder 1.9590.01 nm, Length
29.090.1 nm.
The shape of the distribution function (Fig. 8)
calculated from the scattering data using the pro-
gram GNOM (by D. Svergun, EMBL, Hamburg)
suggests that some non-spherical aggregates are
formed, since in the case of spherical micelles a
Gaussian shaped distribution function would have
been found. These results supported our hypothe-
sis that the found phase might be a lyotropic
cholesteric phase, and in the next step model
calculations (for details, see Section 2.2) have
been performed to simulate the SANS pattern of
the two possible kinds of anisotropic micelles,
disc-like micelles and cylindrical micelles. In Fig.
9 the experimental SANS data are shown, to-
gether with the model fits for the two different
Since this phase was found more or less abun-
dantly in the case of stearyl glycosides with a
disaccharide headgroup, the question arose, as to
whether or not its occurrence is restricted to this
long chain compound. To answer this question,
contact preparations using maltobiosides with a
shorter alkyl chain were performed. In the case of
the tetradecyl-b- -maltobioside 36, a lyotropic
D
cholesteric phase was still found by polarising
microscopy. This phase disappeared, if the te-
tradecyl chain was shortened by two CH2-groups
(dodecyl-b-
D
-maltobioside 35). These compounds
have also been analysed by small-angle neutron

172
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
diffraction (SANS). The SANS data of the te-
tradecyl-b- -maltobioside 36 (Fig. 10 a) again
matched well with the model fits for cylindrical
micelles (the size of the micelles was calculated as
above from the SANS data; length of the cylin-
ders 20.0 nm, radius 2.0 nm), while the SANS
The a cellobioside 12 formed on cooling of the
melted sample the already described monotropic
cubic phase, which was brought into contact with
water. With increasing water concentration, a
lamellar phase formed, followed by another cubic
D
phase,
a columnar phase, and a lyotropic
data of dodecyl-b- -maltobioside 35 do not (Fig.
D
cholesteric phase at the highest water concentra-
tion, and the phase sequence is, therefore, VII, La,
VI, HI, ch. From this contact preparation, it can
be concluded that the thermotropic cubic phase of
the cellobioside 12 should also be of the inverted
type of bicontinuous cubic phases (see also Sec-
tion 3 above), and the same holds true for the
monotropic cubic phases of the lactobiosides 16
and 18, since the reason for the formation of this
monotropic cubic phase — i.e. a linear structure
with an extremely imbalanced ratio between the
hydrophilic and hydrophobic moieties — is the
same. After penetration with water, the lacto-
biosides 16 and 18 formed with increasing water
concentration only a cubic, a columnar and prob-
ably also a lyotropic cholesteric phase, i.e. the
phase sequence was VI, HI, and ch. Within the
cubic phase, a sharp boundary could be seen,
10 b). These data match instead well with the
model fit calculated for ellipsoidal micelles (again
the size of the micelles was calculated from the
SANS data; minor radius 2.0 nm, axis ratio 1.8).
This result, together with the outcome of the
contact preparations, can be interpreted in terms
of a chain length-dependence of the occurrence of
a lyotropic cholesteric phase. This phase can only
be found beyond a minimum chain length, which
in the case of the alkyl-b-D-maltobiosides proved
to be a tetradecyloxy chain. Below this chain
length, only slightly unisotropic ellipsoidal mi-
celles are formed which do not adopt a lyotropic
cholesteric phase. The reason for this chain length
dependence might be seen in constraints to the
formation of unisotropic micelles, due to prob-
lems filling the hydrophobic volume of the an-
isotropic micelles in the case of short alkyl chains.
Fig. 9. SANS data for compounds 8 and 10 together with the model fits; solid lines, model fits for cylindrical micelles; dashed lines,
model fits for disc-like micelles.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
173
Fig. 10. SANS data and model fits for, (A) SANS data for mal-b-OC12H25 (35) together with model fits for cylindrical and
ellipsoidal micelles; (B) SANS data for mal-b-OC14H29 (36) together with model fits for cylindrical and ellipsoidal micelles.
separating two isotropic regions. Since no lamel-
lar phase was formed as in the case of the cel-
lobioside 12, it is impossible to decide whether the
inner isotropic region is of the inverted cubic VII
type or a cubic glass, which has formed out of the
cubic phase on cooling. The formation of a cubic
glass on cooling could also account for the disap-
pearance of the additional lamellar La phase.
Since the boundary of the HI phase towards the
water region was somewhat diffuse in the case of
the lactobiosides, the existence of a lyotropic
cholesteric phase, normally forming a thin weakly
anisotropic band besides the columnar phase,
could not be proved unambiguously by polarising
microscopy. However, this seems very likely.
The b cellobioside 14, which crystallised much

174
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
better than the a cellobioside 12, formed only an
La phase, a HI phase, and a lyotropic cholesteric
phase with water.
with shorter alkyl chains (Fig. 11b). However, the
kind of disaccharide and the nature of the linkage
strongly influences the Tc-value. For example, re-
garding the stereoisomeric stearyl melibiosides and
lactosides, the Tc–value of the latter as well as their
state of order is much higher than that of the former
(Fig. 11b). For the melibiosides, the Tc-values are
38 and 67.5°C in a- and b-linkage (20 and 22),
respectively.
The stearyl-a-melibioside 20 formed with in-
creasing water concentration a lamellar La phase,
followed by a region where crystallisation occurred,
followed by a cubic VI phase, a columnar HI phase,
and a lyotropic cholesteric phase at the highest
water concentration. For the b anomer 22, the same
phase sequence was observed except for the bicon-
tinuous VI phase, which disappeared completely
while the region, where crystallisation occurred,
broadened. The explanation for this behaviour
might again be the greater tendency, mentioned
already, of the compounds with a b-linked aliphatic
chain to crystallise.
On penetration with water, the cubic phase of
dodecyl-a-melibioside 26 developed a columnar
phase followed by two isotropic regions separated
from each other by a sharp border. These isotropic
phases in the contact preparation beyond the HI
phase towards higher water concentration are,
according to the general phase behaviour of am-
phiphiles, supposed to be micellar cubic phases, i.e.
the phase sequence is VI, HI, II, II*, but it is rather
unusual that two of them are found and in the case
of the b anomer, the second discontinuous cubic
phase disappeared. This different behaviour must
be attributed to the different linkage of the aliphatic
chain to the melibiose headgroup, yielding in the
case of the a anomer an even broader banana-
shaped structure than in the case of the b anomer.
3.3. Synchrotron radiation X-ray diffraction
The three-dimensional aggregate structures are
also strongly dependent on the sugar- and linkage-
types. The results for the dodecyl melibiosides (26
and 28) and the dodecyl and tridecyl thiomaltosides
(8 and 10) with short alkyl chains are presented in
Fig. 12. The scattering curves basically consist of
two broad intensity maxima around 0.1 and 0.25–
0.30 nm, which might be interpreted as typical for
unilamellar structures (Worthington and Khare,
1978; Bouwstra et al., 1993). For the maltosides,
above 40°C, sharp reflections occur, which could be
typical for multilamellar aggregation. However, the
‘normal’ bilayer L phase can be excluded, because
in that case, the periodicities should be much higher
(for a maltose-containing ether-linked C14-dialkyl-
glycolipid, Hinz et al. (1991) found values above 6
nm). Therefore, an interdigitation of the alkyl
chains may be assumed similarly as found recently
for stearyl monosaccharides (Vill et al., in press),
for these, however, only in the gel phase.
The samples with a C-18 chain and the different
disaccharides maltose, melibiose, cellobiose, and
lactose as headgroup exhibit a much more complex
structural polymorphism. The broad diffraction
maxima indicate the existence of unilamellar struc-
tures, except for the ß-linked maltobioside 4. The
latter undergoes two phase changes (Fig. 13a). At
5°C, the two reflections at 6.22 and 3.58 nm are in
a ratio of ꢀ3, indicating the absence of a lamellar
phase, but rather the presence of the micellar HI
phase. Above 20°C, i.e. above the temperature of
the ‘pre-transition’ (see Fig. 11), clearly a multil-
amellar structure can be observed. Another struc-
tural change occurs above 60°C, the diffraction
patterns at 80°C expressing a strong increase of the
main peak to 5.59 nm and an occurrence of two new
3.2.2. Fourier-transform infrared spectroscopy
The LblLa alkyl chain melting was determined
IR-spectroscopically via the ns(CH2)-vibration. The
phase transition temperatures of the thio malto-
biosides and melibiosides 8, 10, 26, and 28 with
short lipid chains (C12 and C13) lie in the range
B0–7°C, i.e. there are only slight differences
between the O- and S-containing samples (Fig.
11a). The different wavenumber values correspond-
ing to different states of order in the respective
phase states are noteworthy. This relates in partic-
ular for the melibiosides, which are in the a-linkage
more fluid (higher wavenumbers) than in the ß-link-
age. As should be expected, the Tc-values of the
C18-containing samples are much larger than those

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
175
hibits complex diffraction patterns between 5 and
40°C (Fig. 13b), which may be due to a cubic
structure (e.g. the relations hold 3.59 nm=12.5
nm/ꢀ12, and 2.77 nm=12.5 nm/ꢀ20). Above
Tc=40°C, the structure converts into a different,
possibly unilamellar one. The shape of the broad
diffraction maxima indicates, however, that an-
other unresolvable structure may be superim-
posed. A similar polymorphism like that of the
reflections at 4.55 and 2.75 nm. These numbers,
however, are not interconnected by a particular
numerical relation as found in lamellar or cubic
structures indicating possibly the superposition of
different structures. From Fig. 13a, it becomes
clear that the stereoisomeric maltoside and cel-
lobioside behave quite differently, emphasising
the importance of the kind of anomeric linkage.
The melibiose-containing a-linked sample ex-
Fig. 11. Peak position of the symmetric stretching vibration of the methylene groups ns(CH2) vs. temperature for various, (a)
short-chain disaccharide glycolipids; and (b) stearyl-disaccharide glycolipids.

176
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
Fig. 12. Small-angle X-ray diffraction patterns for various short-chain disaccharide glycolipids in the temperature range 5–80°C.
Water content, 90%.
latter sample is expressed by the cellobiose-con-
taining compound, the only difference is the
higher temperature at which the change into a
mainly unilamellar phase occurs, corresponding
to the higher value of Tc=55°C.
biosides the a-anomer has a reflection at 0.451
nm, whereas that of the b-anomer is centred at
0.420 to 0.424 nm, indicating, in accordance to
the infrared data, a higher order of the latter
anomeric configuration. Table 1 summarises the
results of the infrared spectroscopic and the X-ray
diffraction experiments. In accordance with the
microscopic contact preparations, cel-a-OC18 12
forms a cubic phase, whereas for the lactosides
only lamellar phases are observed in contrast to
the contact preparation. For an understanding, it
should be considered that the X-ray data were
obtained at a much higher (90%) water content
than in the microscopic preparations (see ly-
otropism). The reduction of the water concentra-
tion in X-ray diffraction experiments to 55% gave
results very similar to those obtained at 90%
buffer content, i.e. a lyotropic phase behaviour as
described above should play a role only at water
concentrations below 50%.
The two lactosides exhibit a very similar be-
haviour over the entire temperature range (data
not shown). The patterns might be interpreted as
resulting from lamellar structures — the broad-
ness of the peaks indicates a low number of
lamellae. However, a jump of the peak maximum
to higher values (4.76 and 4.65 nm, respectively)
at Tc=70°C is observed, which does not indicate
a pure LßlLa transition, because this should be
accompanied by a decrease rather than an in-
crease of the periodicity. The latter observation
could be consistent with a transition from a lamel-
lar into a non-lamellar phase as previously found
for lipid A from Salmonella minnesota for the
LlHII transition (Brandenburg et al., 1998).
For some of these glycosides, also wide-angle
X-ray diffraction experiments were performed to
determine the short-range order, i.e. the packing
of the alkyl chains, particularly in their gel phase.
It was found that, for example, for the two meli-
Although the investigated disaccharide head-
groups exclusively consist of a ‘glu–glu’ or ‘glu–
gal’ compound, an enormous diversity of
structural polymorphism (alkyl chain melting and
order, types of aggregate structures) are observed

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
177
dependent of the kind of sugars and of the
Acknowledgements
anomeric linkage to the acyl chain and within the
two sugars. These observations might give an idea
for an understanding of the complex biological
functions of such glycolipids in nature.
We are grateful for financial support from the
Deutsche Forschungsgemeinschaft (SFB 470,
projects B5 and B6).
Fig. 13. Small-angle X-ray diffraction patterns in the temperature range 5–80°C for, (a) the stereoisomers maltoside and cellobioside
in b-linkage; and (b) the two melibioside anomers. Water content, 90%.

178
H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
References
Handbook of Liquid Crystals, vol. 3. Wiley-VCH, Wein-
heim, pp. 341–392.
8
Fischer, E., Helferich, B., 1911. Uber neue synthetische gly-
coside. Liebigs Ann. Chem. 383, 68–91.
Beginn, U., Keinath, S., Moeller, M., 1997. Liq. Cryst. 23,
35–41.
Fischer, S., Fischer, H., Diele, S., Pelzl, G., Jankowski, K.,
Schmidt, R.R., Vill, V., 1994. On the structure of the
thermotropic cubic mesophase. Liq. Cryst. 17, 855–861.
Gabriel, A., Dauvergne, F., 1982. The localization method
used at EMBL. Nucl. Instr. Methods 201, 223–224.
Glatter, O., 1977. Data evaluation in small angle scattering:
calculation of the radial electron density distribution by
means of indirect Fourier transformation. Acta Phys. Aust.
47, 83–102.
Glatter, O., 1992. Mathematical, physical and chemical tables.
In: Wilson, A.J. (Ed.), International Tables for Crytalogra-
phy. Kluwer Academic Publishers, Dordrecht.
Glatter, O., Kratky, O. (Eds.), 1982. Small-Angle X-ray Scat-
tering. Academic Press, London, p. 431.
Blunk, D., Praefke, K., Vill, V., 1998. Amphotropic liquid
crystals. In: Demus, D., Goodby, J., Gray, G.W., Spiess,
H.-W., Vill, V. (Eds.), Handbook of Liquid Crystals, vol.
3. Wiley-VCH, Weinheim, pp. 305–340.
Bo¨cker, T., Thiem, J., 1989. Synthese und eigenschaften von
kohlenhydrattensiden. Tenside Surf. Det. 26, 318–324.
Borisch, K., Diele, S., Go¨ring, P., Mu¨ller, H., Tschierske, C.,
1997. Amphiphilic N-benzoyl-1-amino-1-deoxy-D-glucitol
derivatives forming thermotropic lamellar, columnar and
different types of cubic mesophases. Liq. Cryst. 22, 427–
443.
Bouligand, Y., 1998. Defects and textures. In: Demus, D.,
Goodby, J., Gray, G.W., Spiess, H.-W., Vill, V. (Eds.),
Handbook of Liquid Crystals, vol. 1. Wiley-VCH, Wein-
heim, pp. 406–453.
Boulin, C., Kempf, R., Koch, M.H.J., McLaughlin, S.M.,
1986. Data appraisal, evaluation and display for synchro-
tron radiation experiments: hardware and software. Nucl.
Instr. Methods A249, 399–407.
Hori, R., 1958. Alkylderivatives of carbohydrates. II. Synthe-
sis of alkyl-b-cellobiosides. Yakugaku Zasshi 78, 999–
1002.
Israelachvili, J.N., 1991. Intermolecular and Surface Forces.
Academic Press, London.
Jeffrey, G.A., 1984. Carbohydrate liquid crystals. Mol. Cryst.
Liq. Cryst. 110, 221–237.
Jeffrey, G.A., Wingert, L.M., 1992. Carbohydrate liquid crys-
tals. Liq. Cryst. 12, 179–202.
Koch, R., 1884. Die aetiologie der tuberkulose. Mitt. K.
Gesundheitsamt. 2, 1–88.
Koynova, R., Caffrey, M., 1994. Phases and phase transitions
of the glycoglycerolipids. Chem. Phys. Lipids 69, 181–207.
Lindblom, R., Rilfors, R., 1989. Cubic phases and isotropic
structures formed by membrane lipids — possible biologi-
cal relevance. Biochim. Biophys. Acta 988, 221–256.
Luzzati, V., Gulik, A., Gulik-Krzywicki, T., Tardieu, A., 1986.
Lipid polymorphism revisited: structural aspects and bio-
logical implications. In: Lipids and Membranes: Past,
Present, and Future. Elsevier, Amsterdam, pp. 137–151.
Mannock, D.A., Lewis, R.N.A.H., McElhaney, R.N., 1987.
An improved procedure for the preparation of 1,2-di-O-
Boullanger, P., 1998. Amphiphilic carbohydrates as a tool for
molecular recognition in organized systems. In: Drigenz,
H., Thiem, J. (Eds.), Topics in Current Chemistry, vol.
187. Springer, Berlin Heidelberg, pp. 275–312.
Brandenburg, K., Koch, M.H.J., Seydel, U., 1990. Phase
diagram of lipid A from Salmonella minnesota and Es-
cherichia coli rough mutant lipopolysaccharide. J. Struct.
Biol. 105, 11–21.
Brandenburg, K., Richter, W., Koch, M.H.J., Meyer, H.W.,
Seydel, U., 1998. Characterization of the nonlamellar cubic
and HII structures of lipid A from Salmonella enterica
serovar minnesota by X-ray diffraction and freeze-fracture
electron microscopy. Chem. Phys. Lipids 91, 53–69.
Casal, H.L., Mantsch, H.H., 1984. Polymorphic phase be-
haviour of phospholipid membranes studied by infrared
spectrosopy. Biochim. Biophys. Acta 779, 381–401.
Curatolo, W., 1987. The physical properties of glycolipids.
Biochim. Biophys. Acta 906, 111–136.
acyl-3-O-(b-D-glucopyranosyl)-sn-glycerols. Chem. Phys.
Lipids 43, 113–127.
Mannock, D.A., Lewis, R.N.A.H., McElhaney, R.N., 1990.
The chemical synthesis and physical characterisation of
Dahlhoff, W.V., 1987. Amphiphilic carbohydrate-based meso-
gens 2, aldose di-n-alkyl dithioacetals: novel double-tailed
thermotropic liquid crystals. Z. Naturforsch. 42b, 661–
662.
1,2-di-O-acyl-3-O-(a-D-glucopyranosyl)-sn-glycerols,
an
important class of membrane glycolipids. Chem. Phys.
Lipids 55, 309–321.
Dahlhoff, W.V., 1990. Synthesis of a series of alkyl 1-tio-
D-
glucopyranosides and their regioselective reductions to 1-
Niori, T., Sekine, T., Watanabe, J., Furukawa, T., Takezoe,
H., 1996. J. Mater. Chem. 6, 1231–1233.
alkylthio-1-deoxy-
1025–1027.
D-glucitols. Liebigs Ann. Chem.,
Pedersen, J.S., 1997. Analysis of small-angle scattering data
from colloids and polymer solutions: modelling and least-
squares fitting. Adv. Colloid Interface Sci. 70, 171–210.
Prade, H., Miethchen, R., Vill, V., 1995. Thermotrop flu¨s-
sigkristalline kohlenhydrat-amphiphile. J. Prakt. Chem.
337, 427–440.
Eckert, A., Kohne, B., Praefcke, K., 1987. Flu¨ssigkristalline
kohlenhydratdithioacetale. Z. Naturforsch. 43b, 878–888.
Ellens, H., Siegel, D.P., Alford, D., Yeagle, P.L., Boni, L., Lis,
J., Quinn, P.J., Bentz, J., 1989. Membrane fusion and
inverted phases. Biochemistry 28, 3692–3703.
Fairhurst, C.E., Fuller, S., Gray, J., Holmes, M.C., Tiddy,
G.J., 1998. Lyotropic surfactant liquid crystals. In: Demus,
D., Goodby, J., Gray, G.W., Spiess, H.-W., Vill, V. (Eds.),
Praefcke, K., Kohne, B., Eckert, A., Hempel, J., 1990.
Inosose-flu¨ssigkristalle. Z. Naturforsch. 45b, 1084–1090.

H.M. 6on Minden et al. / Chemistry and Physics of Lipids 106 (2000) 157–179
179
Rosevar, P., VanAken, T., Baxter, J., Ferguson-Miller, 1980.
cose dialkyl acetals and dodecyl
Chem. Ber. 127, 1065–1068.
D
-glucofuranosides.
Alkyl glycoside detergents: a simple synthesis and their
effects on kinetic and physical properties of cytochrome c
oxidase. Biochim. Biophys. Acta 19, 4108–4115.
Seddon, J.M., 1990. Structure of the inverted hexagonal (HII)
phase, and non-lamellar phase transitions of lipids.
Biochim. Biophys. Acta 1031, 1–69.
Van Doren, H.A., Wingert, L.M., 1994. The relationship
between the molecular structure of polyhydroxy am-
phiphiles and their aggregation behaviour in water; I. The
contact preparation method as a tool for empirical studies.
Recl. Trav. Chim. Pays-Bas 113, 260–265.
Seydel, U., Labischinski, H., Kastowsky, M., Brandenburg,
K., 1993. Phase behaviour, supramolecular structure, and
molecular conformation of lipopolysaccharide. Immunobi-
ology 187, 191–211.
Siegel, D.P, 1986a. Inverted micellar intermediates and the
transition between lamellar, cubic and inverted hexagonal
lipid phases; I. Mechanism of the La-HII phase transition.
Biophys. J. 49, 1155–1170.
Siegel, D.P, 1986b. Inverted micellar intermediates and the
transition between lamellar, cubic and inverted hexagonal
lipid phases; II. Implications for membrane–membrane
interactions and membrane-fusion. Biophys. J. 49, 1171–
11783.
Siegel, D.P, 1986c. Inverted micellar intermediates and the
transition between lamellar, cubic and inverted hexagonal
lipid phases; III. Isotropic and inverted cubic state forma-
tion via intermediates in transitions between La and HII
phases. Chem. Phys. Lipids 42, 279–301.
Siegel, D.P., Banschbach, J., Alford, J., Ellens, H., Lis, L.J.,
Quinn, P.J., Yeagle, P.L., Bentz, J., 1989. Physiological
levels of diacylglycerols in phospholipid membranes induce
membrane fusion and stabilize inverted phases. Biochem-
istry 28, 3703–3709.
Stuhrmann, H.B., 1989. Small-angle scattering of X-rays. In:
Progress Crystal Growth and Characteristics, vol. 18. Perg-
amon Press, Oxford, pp. 1–18.
Stuhrmann, H.B., Burkhardt, N., Dietrich, G., Ju¨nemann, R.,
Meerwinck, W., Schmitt, M., Wadzack, J., Willumeit, R.,
Zhao, J., Nierhaus, K.H., 1995. Proton- and deuteron-spin
targets in biological structure research. Nucl. Instr. Meth-
ods A356, 133–137.
Van Doren, H.A., Buma, T.J., Kellog, R.M., Wynberg, H.,
1988. Carbohydrate liquid crystals even at ambient temper-
ature. J. Chem. Soc. Chem. Commun., 460–462.
Van Doren, H.A., van der Geest, R., Kellogg, M.K., Wyn-
berg, H., 1989. Synthesis and liquid crystalline properties
of the n-alkyl 1-thio-a-D-glucopyranosides,
a new ho-
mologous series of carbohydrate mesogens. Carbohydr.
Res. 194, 71–77.
Van Doren, H.A., van der Geest, R., Kellogg, M.K., Wyn-
berg, H., 1990. A comparative study of the liquid-crys-
talline behavior of three homologous series of
1,2-propanediol derivatives. Recl. Trav. Chim. Pays-Bas
109, 197–203.
Vill, V., 1999. LiqCryst 3.3-database of liquid crystalline com-
pounds for personal computers. Fujitsu Kyushu Systems
(FQS) Ltd., Fukuoka, Japan; LCI Publisher, Hamburg,
1999. http://liqcryst.chemie.uni-hamburg.de
Vill, V., Bo¨cker, T., Thiem, J., Fischer, F., 1989. Studies on
liquid crystalline glycosides. Liq. Cryst. 6, 349–356.
Vill, V., Kelkenberg, H., Thiem, J., 1992. Liquid-crystalline
glucamin derivatives. Liq. Cryst. 11, 459–467.
Vill, V., v. Minden, H. M., Koch, M. H. J., Seydel, U.,
Brandenburg, K., 1999. Thermotropic and lyotropic prop-
erties of long chain alkyl glycopyranosides. Part I:
monosaccharide headgroups. Chem. Phys. Lipids, in press.
Vogel, C., Jeschke, U., Vill, V., Fischer, H., 1992. D-Galactur-
onsa¨urederivate mit flu¨ssigkristallinen eigenschaften.
Liebigs Ann. Chem., 1171–1177.
Wignall, G.D., Bates, F.S., 1986. Absolute calibration of
small-angle neutron scattering data. J. Appl. Cryst. 20,
28–40.
Tietze, L.F., Bo¨ge, K., Vill, V., 1994. Liquid-crystalline D-glu-
.