Thermotropic and lyotropic properties of long chain alkyl glycopyranosidesPart I: Monosaccharide headgroups

Article abstract of DOI:10.1016/S0009-3084(99)00119-X

A systematic structure variation of a classical amphiphile (dodecyl-β-D-glucopyranoside) is performed, demonstrating the influence of anomeric linkage, configuration, ring size and flexibility as well as electric charges on the mesophase behaviour. In addition, we have investigated the thermotropic and lyotropic properties of some long chain alkyl glycosides with monosaccharide headgroups. The thermotropism was measured with polarizing microscopy and differential scanning calorimetry, and additionally the lyotropism with FTIR spectroscopy and X-ray diffraction. Copyright (C) 2000 Elsevier Science Ireland Ltd.

Full text of DOI:10.1016/S0009-3084(99)00119-X

Chemistry and Physics of Lipids
104 (2000) 75–91
www.elsevier.com/locate/chemphyslip
Thermotropic and lyotropic properties of long chain alkyl
glycopyranosides
Part I: monosaccharide headgroups
V. Vill a,*, H.M. von Minden a, M.H.J. Koch b, U. Seydel c, K. Brandenburg c
a Institut fu¨r Organische Chemie, Uni6ersita¨t Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg, Germany
b European Molecular Biology Laboratory, c/o DESY, Notkestr. 85, D-22603 Hamburg, Germany
c Forschungszentrum Borstel, Laborgruppe Biophysik, Parkallee 10, 23845 Borstel, Germany
Received 4 May 1999; received in revised form 6 September 1999; accepted 21 September 1999
Abstract
A systematic structure variation of a classical amphiphile (dodecyl-b-D-glucopyranoside) is performed, demonstrat-
ing the influence of anomeric linkage, configuration, ring size and flexibility as well as electric charges on the
mesophase behaviour. In addition, we have investigated the thermotropic and lyotropic properties of some long chain
alkyl glycosides with monosaccharide headgroups. The thermotropism was measured with polarizing microscopy and
differential scanning calorimetry, and additionally the lyotropism with FTIR spectroscopy and X-ray diffraction.
© 2000 Elsevier Science Ireland Ltd. All rights reserved.
Keywords: Alkyl glycopyranosides; Thermotropic liquid crystals; Lyotropic liquid crystals; Phase transitions; Glycolipids
motropic liquid crystalline phases in their pure
state upon heating and lyotropic liquid crystalline
1. Introduction
phases upon addition of a solvent. The first obser-
The observation of a double melting of certain
vation of lyotropic behaviour is also related to
long chain alkyl glucopyranosides, e.g. hexadecyl-
studies on alkylated carbohydrates: while
b-D-glucopyranoside (Fig. 1) by Fischer and
analysing the extracts from tuberculosis bacteria
Koch (1884) observed unusual optical textures of
their aqueous dispersions.
Helferich (1911) was the first indication of ther-
motropic liquid crystalline properties in am-
phiphilic carbohydrates.
The driving force for the mesophase formation
in the case of the amphiphilic molecules is a micro
phase separation leading to an aggregate structure
with separated regions for the lipophilic and hy-
drophilic molecular moieties, enabling the mainte-
nance of the van der Waals interaction in the
These amphotropic molecules form both ther-
* Corresponding author. Tel.: +49-40-42838-4269; fax: +
49-40-42838-4325.
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(99)00119-X

76
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
Fig. 1. Hexadecyl-b-
D-glucopyranoside.
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) because of their interesting proper-
ties as surfactants (Vill et al., 1986; Bo¨cker and
Thiem, 1989; Boullanger, 1998). These com-
pounds dissolve in water, whereas the long chain
glycosides will only swell.
We synthesised long-chain alkyl glycopyra-
nosides (in particular stearyl glycosides) which
can be incorporated in biomembranes, and
changed the sugar headgroup systematically to
elucidate the structure–property relationships
governing the occurrence of the above mentioned
polymorphism. In the case of a monosaccharide
headgroup we observed mainly SA phases,
whereas a broad range of phases could be ob-
served with compounds having a disaccharide
headgroup. In the first part of this paper we
would like to report on our results obtained with
long chain-alkyl monoglycosides as well as to
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, while simple amphiphiles such
as the one shown in Fig. 1 normally exhibit
thermotropically only an SA phase.
Deviation from the above mentioned vertical
line may also be seen upon the addition of sol-
vents, the most common of which is water, be-
cause 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; Ellens et al., 1989;
Lindblom and Rilfors, 1989; Siegel et al., 1989).
The lamellar phase with its multibilayer structure
has long been well known to the biologist, but
there is much evidence today, that the bicontinu-
ous cubic phase may play an important role in
processes like membrane fusion.
Fig. 2. Principal phase behaviour of amphiphilic compounds.

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
77
2.1.2. Stearyl-2,3,4,6-tetra-O-acetyl-h-
pyranoside 1
General reaction conditions (Vill et al., 1986)
D-gluco-
using penta-O-acetyl-b-D-glucopyranose and
stearyl alcohol were used.
1H NMR (400 MHz, CDCl3+TMS): l=5.48
(dd, 1H, H-3), 5.05 (dd, 1H, H-4), 5.06 (d, 1H,
H-1), 4.85 (dd, 1H, H-2), 4.26 (dd,1H, H-6a),
4.09 (dd, 1H, H-6b), 4.02 (ddd, 1H, H-5), 3.68
(dt, 1H, H-aa), 3.42 (dt, 1H, H-ab), 2.12, 2.06,
2.03, 2.01 (je s, 3H, OAc), 1.54–1.64 (m, 2H,
b-CH2), 1.20–1.38 (m, 30H, –CH2–), 0.88
3
(t, 3H, –CH3); 3J1,2=3.6, 3J2,3=10.2, J3,4
=
=
3
3
3
2
9.7, J4,5=10.2, J5,6a=4.6, J5,6b=2.0, J6a,6b
2
3
3
12.2, Jaa,ab=9.7, Jaa,b-CH =6.6, Jab,b-CH =6.6
2
2
Hz.
2.1.3. Stearyl-h-
D
-glucopyranoside 2
Stearyl-2,3,4,6-tetra-O-acetyl-a-
D
-glucopyra-
noside 1 was deprotected using the general reac-
tion conditions (Vill et al., 1986).
1H NMR (400 MHz, d4-MeOH): l=4.81 (d,
1H, H-1), 3.84 (dd, 1H, H-6a), 3.78 (dt, 1H,
H-aa), 3.71 (dd, 1H, H-6b,), 3.68 (dd, 1H, H-3),
3.61 (ddd, 1H, H-5), 3.48 (dt, 1H, H-ab), 3.42
(dd, 1H, H-2), 3.33 (dd, 1H, H-4), 1.68 (mc, 2H,
b-CH2), 1.24–1.50 (m, 30H, –CH2–), 0.95 (t,
3H, –CH3); 3J1,2=3.6, 3J2,3=9.7, 3J3,4=10.2,
3J4,5=9.7, 3J5,6a=2.5, 3J5,6b=5.6, 2J6a,6b=11.7,
Fig. 3. Synthesis of alkyl glycopyranosides.
discuss a homologous series of dodecyl mono-
glycosides.
2. Materials and methods
3
3
2Jaa,ab=9.7, Jaa,b-CH =6.6, Jab,b-CH =6.6
2
2
2.1. Materials
2.1.4. Stearyl-2,3,4,6-tetra-O-acetyl-i-
pyranoside 3
D
-gluco-
2.1.1. General reaction conditions
The alkyl glycosides have been synthesised as
shown in Fig. 3 for the glucosides using the
method described by Vill et al. (1986), however,
boron trifluoride diethyl etherate was used in-
stead of tin tetrachloride as the Lewis acid for
the synthesis of the b-anomers. Furthermore, no
ion exchange chromatography was necessary to
separate the anomers, since silica gel chromatog-
raphy (light petroleum (b.p. 60–70°C)–ethyl ac-
etate 4:1) was already successful.
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b-D-glucopyranose and
stearyl alcohol were used.
1H NMR (400 MHz, CDCl3+TMS): l=5.21
(dd, 1H, H-3), 5.09 (dd, 1H, H-4), 4.99 (dd, 1H,
H-2), 4.49 (d, 1H, H-1), 4.27 (dd, 1H, H-6a),
4.14 (dd, 1H, H-6b), 3.87 (dt, 1H, H-aa), 3.69
(ddd, 1H, H-5), 3.47 (dt, 1H, H-ab), 2.09, 2.04,
2.03. 2.01 (je s, 3H, OAc), 1.48–1.63 (m, 2H,
b-CH2), 1.17–1.34 (m, 30H, –CH2–), 0.88 (t,
3H, –CH3); 3J1,2=8.1, 3J2,3=9.7, 3J3,4=9.7,
3J4,5=9.7, 3J5,6a=4.6, 3J5,6b=2.6, 2J6a,6b=12.2,
The synthesis of the compounds A–K in Fig. 4
is described elsewhere (Vill et al., 1986, 1991;
Bo¨cker et al., 1992; Stanger et al., 1994; Tietze et
al., 1994).
3
3
2Jaa,ab=9.7, Jaa,b-CH =6.7, Jab,b-CH =6.9 Hz.
2
2

78
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
Fig. 4. Structural modification of a dodecyl-b-D-glucopyranoside. Cr, crystalline; SA, smectic A; SB, smectic B; I, isotropic melting;
the transition temperatures are given in °C.

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
79
2.1.5. Stearyl-i-
Stearyl-2,3,4,6-tetra-O-acetyl-b-
D
-glucopyranoside 4
-glucopyra-
using penta-O-acetyl-b-
D
-glucopyranose and
D
oleoyl alcohol were used.
1H NMR (400 MHz, CDCl3+TMS): l=5.48
(dd, 1H, H-3), 5.05 (dd, 1H, H-4), 5.06 (d, 1H,
H-1), 4.85 (dd, 1H, H-2), 4.26 (dd,1H, H-6a), 4.09
(dd, 1H, H-6b), 4.02 (ddd, 1H, H-5), 3.48 (dd,
1H, H-aa), 3.21 (dd, 1H, H-ab), 2.12, 2.06, 2.03,
2.01 (je s, 3H, OAc), 1.66–1.80 (m, 4H, cyclo-
hexyl), 1.44–159 (m, 2H, cyclohexyl), 1.08–1.34
(m, 14H, –CH2–), 0.80–0.97 (m, 7H, –CH3, cy-
clohexyl); 3J1,2=3.6, 3J2,3=10.2, 3J3,4=9.7,
noside 3 was deprotected using the general reac-
tion conditions (Vill et al., 1986).
1H NMR (400 MHz, d4-MeOH): l=4.28 (d,
1H, H-1), 3.94 (dt, 1H, H-aa), 3.90 (dd, 1H,
H-6a), 3.70 (dd, 1H, H-6b), 3.57 (dt, 1H, H-ab),
3.26–3.42 (m, 3H, H-3, H-4, H-5), 3.2 (dd, 1H,
H-2), 1.66 (mc, 2H, b-CH2), 1.26–1.46 (m, 30H,
3
3
–CH2–), 0.94 (t, 3H, –CH3); J1,2=7.6, J2,3
=
9.2,
3J5,6a=2.0,
3J5,6b=5.1,
2J6a,6b=11.7,
3
3
2Jaa,ab=9.7, Jaa,b-CH =7.1, Jab,b-CH =6.6 Hz.
3
3
2
3J4,5=10.2, J5,6a=4.6, J5,6b=2.0, J6a,6b=12.2,
2
2
3
3
2Jaa,ab=9.7, Jaa,b-CH =6.6, Jab,b-CH =6.1 Hz.
2
2
2.1.6. Oleyl-2,3,4,6-tetra-O-acetyl-i-
D-gluco-
pyranoside 5
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b-D-glucopyranose and
oleoyl alcohol were used.
2.1.9. 4%-Octylcyclohexylmethyl-h-
oside 8
D
-glucopyran-
4%-Octylcyclohexylmethyl-2,3,4,6-tetra-O-acetyl-
-glucopyranoside 7 was deprotected using the
a-D
1H NMR (400 MHz, CDCl3+TMS): l=
5.28–5.41 (m, 2H, H-olefinic), 5.21 (dd, 1H, H-3),
5.09 (dd, 1H, H-4),4.99 (dd, 1H, H-2), 4.49 (d,
1H, H-1), 4.27 (dd,1H, H-6a), 4.14 (dd, 1H, H-
6b), 3.87 (dt, 1H, H-aa), 3.69 (ddd, 1H, H-5), 3.47
(dt, 1H, H-ab), 2.09, 2.04, 2.03. 2.01 (je s, 3H,
OAc), 1.96–2.05 (m, 4H, –CH2– allylic), 1.48–
1.64 (m, 2H, b-CH2), 1.20–1.37 (m, 22H, –CH2–),
general reaction conditions (Vill et al., 1986).
1H NMR (400 MHz, d4-MeOH): l=4.78 (d,
1H, H-1), 3.83 (dd, 1H, H-6a), 3.70 (dd, 1H,
H-6b,), 3.68 (dd, 1H, H-3), 3.60 (ddd, 1H, H-5),
3.56 (dd, 1H, H-aa), 3.42 (dd, 1H, H-2), 3.25–
3.43 (m, 1H, H-4, H-ab), 1.77–1.98 (m, 4H, cy-
clohexyl), 1.65 (mc, 1H, cyclohexyl), 1.25–1.41
(m, 14H, –CH2–), 1.17–1.25 (m, 1H, cyclohexyl),
3
3
3
0.88 (t, 3H, –CH3); J1,2=8.1, J2,3=9.7, J3,4
=
=
=
9.7, 3J4,5=9.7, 3J5,6a=4.6, 3J5,6b=2.6, J6a,6b
2
3
0.88–1.10 (m, 7H, –CH3, cyclohexyl); J1,2=4.1,
2
3
3
3J2,3=9.7, 3J3,4=9.2, 3J4,5=9.7, 3J5,6a=2.5,
12.2, Jaa,ab=9.7, Jaa,b-CH =6.6, Jab,b-CH
2
2
3J5,6b=5.6,
2J6a,6b=11.7,
2Jaa,ab=9.1,
6.9 Hz.
3Jaa,cyclohexyl=7.1 Hz.
2.1.7. Oleyl-i-
D
-glucopyranoside 6
Oleyl - 2,3,4,6 - tetra - O - acetyl - b -
D
- glucopyra-
2.1.10. 4%-Octylcyclohexylmethyl-2,3,4,6-tetra-O-
acetyl-i- -glucopyranoside 9
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b-D-glucopyranose and
oleoyl alcohol were used.
noside 5 was deprotected using the general reac-
tion conditions (Vill et al., 1986).
D
1H NMR (400 MHz, d4-MeOH): l=5.34–5.45
(m, 2H, H-olefinic), 4.28 (d, 1H, H-1), 3.94 (dt,
1H, H-aa), 3.90 (dd, 1H, H-6a), 3.70 (dd, 1H,
H-6b), 3.57 (dt, 1H, H-ab), 3.25–3.41 (m, 3H,
H-3, H-4, H-5), 3.21 (dd, 1H, H-2), 1.97–2.13 (m,
4H, H-allylic), 1.66 (mc, 2H, b-CH2), 1.28–1.47
1H NMR (400 MHz, CDCl3+TMS): l=5.20
(dd, 1H, H-3), 5.08 (dd, 1H, H-4), 4.99 (dd, 1H,
H-2), 4.46 (d, 1H, H-1), 4.26 (dd, 1H, H-6a), 4.13
(dd, 1H, H-6b), 3.24 (dd, 1H, H-aa), 3.70 (dd,
1H, H-ab), 3.67 (ddd, 1H, H-5), 2.08, 2.03, 2.02,
2.01 (je s, 3H, OAc), 1.66–1.80 (m, 4H, cyclo-
hexyl), 1.44–159 (m, 2H, cyclohexyl), 1.08–1.34
(m, 14H, –CH2–), 0.80–0.97 (m, 7H, –CH3, cy-
clohexyl); 3J1,2=8.1, 3J2,3=9.7, 3J3,4=9.7,
3J4,5=9.7, 3J5,6a=4.6, 3J5,6b=2.6, 2J6a,6b=12.2,
3
(m, 30H, –CH2–), 0.94 (t, 3H, –CH3); J1,2=7.6,
3J2,3=9.2, 3J5,6a=2.0, 3J5,6b=5.6, 2J6a,6b=12.2,
3
3
2Jaa,ab=9.7, Jaa,b-CH =6.6, Jab,b-CH =6.6 Hz.
2
2
2.1.8. 4%-Octylcyclohexylmethyl-2,3,4,6-tetra-O-
acetyl-h- -glucopyranoside 7
General reaction conditions (Vill et al., 1986)
D

80
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
3
3
2Jaa,ab=9.7, Ja,cyclohexyl=7.1, Jb,cyclohexyl=6.1
Hz.
using penta-O-acetyl-b-D-glucopyranose and
oleoyl alcohol were used.
1H NMR (400 MHz, CDCl3+TMS): l=5.41
(dd, 1H, H-4), 5.23 (dd, 1H, H-2), 5.04 (dd, 1H,
H-3), 4.47 (d, 1H, H-1), 4.21 (dd,1H, H-6a), 4.15
(dd, 1H, H-6b), 3.88–3.95 (m, 2H, H-aa, H-5),
3.49 (dt, 1H, H-ab), 2.17, 2.01. (je s, 3H, OAc),
2.07 (s, 6H, OAc), 1.51–1.67 (m, 2H, b-CH2),
1.20–1.44 (m, 30H, –CH2–), 0.90 (t, 3H, –CH3);
3J1,2=8.2, 3J2,3=10.2, 3J3,4=3.6, 3J4,5=1.4,
3J5,6a=6.3, 3J5,6b=6.9, 2J6a,6b=11.4, 2Jaa,ab=9.6,
2.1.11. 4%-Octylcyclohexylmethyl-i-
oside 10
D-glucopyran-
4%-Octylcyclohexylmethyl-2,3,4,6-tetra-O-acetyl-
-glucopyranoside 9 was deprotected using the
b-
D
general reaction conditions (Vill et al., 1986).
1H NMR (400 MHz, d4-MeOH): l=4.26 (d,
1H, H-1), 3.90 (dd, 1H, H-6a), 3.75 (dd 1H,
H-aa), 3.70 (dd, 1H, H-6b), 3.25–3.41 (m, 4H,
H-3, H-4, H-5, H-ab), 3.21 (dd, 1H, H-2), 1.77–
1.95 (m, 4H, cyclohexyl), 1.61 (mc, 1H, cyclo-
hexyl), 1.25–1.41 (m, 14H, –CH2–), 1.17–1.25
(m, 1H, cyclohexyl), 0.87–1.08 (m, 7H, –CH3,
cyclohexyl); 3J1,2=8.1, 3J2,3=9.2, 3J5,6a=2.0,
3Jab,b-CH =6.9 Hz.
2
2.1.15. Stearyl-i-
Stearyl-2,3,4,6-tetra-O-acetyl-b-
D
-galactopyranoside 14
-galactopyra-
D
noside 13 was deprotected using the general reac-
tion conditions (Vill et al., 1986).
3J5,6b=5.1,
2J6a,6b=11.7,
2Jaa,ab=9.2,
3Jaa,cyclohexyl=6.6 Hz.
1H NMR (400 MHz, d4-MeOH): l=4.25 (d,
1H, H-1), 3.94 (dt, 1H, H-aa), 3.87 (dd, 1H, H-4),
3.80 (dd, 1H, H-6a), 3.76 (dd, 1H, H-6b,), 3.58
(dt, 1H, H-ab), 3.51–3.57 (m, 2H, H-2, H-5), 3.49
(dd, 1H, H-3), 1.67 (mc, 2H, b-CH2), 1.28–1.46
2.1.12. Stearyl-2,3,4,6-tetra-O-acetyl-h-
ctopyranoside 11
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b-D-glucopyranose and
oleoyl alcohol were used.
D-gala-
3
(m, 30H, –CH2–), 0.95 (t, 3H, –CH3); J1,2=7.1,
3J2,3=9.7, 3J3,4=3.1, 3J4,5=1.0, 3J5,6a=6.6,
1H NMR (400 MHz, CDCl3+TMS): l=5.46
(dd, 1H, H-4), 5.35 (dd, 1H, H-3), 5.08-5.14 (m,
2H, H-1, H-2), 4.22 (ddd, 1H, H-5), 4.11 (dd,1H,
H-6a), 4.08 (dd, 1H, H-6b), 3.68 (dt, 1H, H-aa),
3.42 (dt, 1H, H-ab), 2.14, 2.07, 2.04, 1.99 (je s,
3H, OAc), 1.54–1.64 (m, 2H, b-CH2), 1.20–1.38
3J5,6b=5.6, 2J6a,6b=11.2, Jaa,ab=9.2, Jaa,b-
2
3
3
CH2=6.6, Jab,b-CH =6.6 Hz.
2
2.1.16. Oleyl-2,3,4,6-tetra-O-acetyl-i-D-galacto-
pyranoside 15
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b- -galactopyranose and
oleoyl alcohol were used.
3
(m, 30H, –CH2–), 0.88 (t, 3H, –CH3); J3,4=3.6,
D
3J4,5=1.0, 3J5,6a=6.1, 3J5,6b=7.1, 2J6a,6b=11.2,
3
3
2Jaa,ab=9.7, Jaa,b-CH =6.7, Jab,b-CH =6.7 Hz.
1H NMR (400 MHz, CDCl3+TMS): l=5.39
(dd, 1H, H-4), 5.28–5.38 (m, 2H, H-olefinic), 5.21
(dd, 1H, H-2), 5.02 (dd, 1H, H-3), 4.45 (d, 1H,
H-1), 4.19 (dd, 1H, H-6a), 4.13 (dd,1H, H-6b),
3.85–3.93 (m, 2H, H-5, H-aa), 3.47 (dt, 1H, H-
ab), 2.15 (s, 3H, OAc), 2.05 (s, 6H, OAc), 1.98 (s,
3H, OAc), 1.93–2.07 (m, 4H, –CH2– allylic),
1.48–1.64 (m, 2H, b-CH2), 1.20–1.37 (m, 22H,
2
2
2.1.13. Stearyl-h-
Stearyl-2,3,4,6-tetra-O-acetyl-a-
D
-galactopyranoside 12
-galactopyra-
D
noside 11 was deprotected using the general reac-
tion conditions (Vill et al., 1986).
1H NMR (400 MHz, d4-MeOH): l=4.84 (d,
1H, H-1), 3.93 (dd, 1H, H-4), 3.85 (ddd, 1H,
H-5), 3.71–3.83 (m, 5H, H-2, H-3, H-6a, H-6b,
H-aa), 3.48 (dt, 1H, H-ab), 1.62–1.75 ( m, 2H,
b-CH2), 1.29–1.46 (m, 30H, –CH2–), 0.95 (t, 3H,
3
3
–CH2–), 0.88 (t, 3H, –CH3); J1,2=8.1, J2,3
=
=
10.2, 3J3,4=3.1, 3J4,5=1.0, 3J5,6a=6.6, J5,6b
3
3
3
3
3
2
2
3
–CH3); J1,2=3.6, J3,4=3.1, J4,5=1.5, J5,6a
=
7.1, J6a,6b=11.2, Jaa,ab=9.7, Jab,b-CH =6.6 Hz.
2
3
2
3
6.1, J5,6b=6.1, Jaa,ab=9.7, Jab,b-CH =6.6 Hz.
2
2.1.17. Oleyl-i-
Oleoyl-2,3,4,6-tetra-O-acetyl-b-
noside 15 was deprotected using the general reac-
D
-galactopyranoside 16
-galactopyra-
2.1.14. Stearyl-2,3,4,6-tetra-O-acetyl-i-
galactopyranoside 13
D
-
D
General reaction conditions (Vill et al., 1986)
tion conditions (Vill et al., 1986).

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
81
1H NMR (400 MHz, d4-MeOH): l=5.28-5.46
(m, 2H, H-olefinic), 4.24 (d, 1H, H-1), 3.93 (dt,
1H, H-aa), 3.87 (dd, 1H, H-4), 3.79 (dd, 1H,
H-6a), 3.76 (dd, 1H, H-6b), 3.5–61 (m, 3H, H-2,
H-5, H-ab), 3.49 (dd, 1H, H-3), 1.96–2.14 (m,
4H, H-allylic), 1.66 (mc, 2H, b-CH2), 1.27–1.46
tations were recorded using a Perkin-Elmer 241
polarimeter. NMR spectra (1H: 400 MHz) were
recorded on a Bruker AMX-400 spectrometer
(mc=centred multiplet). An Olympus BH optical
polarizing microscope equipped with a Mettler FP
82 hot stage and a Mettler FP 80 central proces-
sor was used to identify thermal transitions and
characterise anisotropic textures.
3
(m, 30H, –CH2–), 0.94 (t, 3H, –CH3); J1,2=7.1,
3J2,3=9.7, 3J3,4=3.1, 3J4,5=1.0, 3J5,6a=6.6,
2
3
3J5,6b=5.6, 2J6a,6b=11.5, Jaa,ab=9.7, Jaa,b-
The DSC measurements were performed on a
Mettler DSC 27HP with a Mettler 4000 central
processor.
CH2=6.6 Hz.
For the contact preparation a small amount of
sample was placed on a microscope slide and
covered with a cover glass before heating to the
SA phase. Afterwards a small amount of water
was placed on the slide at the edge of the cover
glass. As soon as the water had moved under the
cover glass and the sample was completely sur-
rounded with water it was placed again for some
seconds on the hot stage at a temperature of
100–120°C and the phase behaviour was investi-
gated immediately afterwards by polarizing
microscopy.
2.1.18. Nonan-1,9-diyl-bis(2%,3%,4%,6%-tetra-O-
acetyl-i-
General reaction conditions (Vill et al., 1986)
using penta-O-acetyl-b- -glucopyranose and 1,9-
nonanedithiol were used.
D
-thioglucopyranoside) 17
D
1H NMR (400 MHz, CDCl3+TMS): l=5.22
(dd, 2H, H-3, H-3%), 5.09 (dd, 2H, H-4, H-4%), 5.03
(dd, 2H, H-2, H-2%), 4.48 (d, 2H, H-1, H-1%), 4.25
(dd, 2H, H-6a, H-6a%), 4.14 (dd, 2H, H-6b, H-6b%),
3.91 (dd, 2H, H-5, H-5%), 2.60–2.74 (m, 4H, a-
CH2, a%-CH2), 2.08, 2.06, 2.03, 2.01 (je s, 6H,
OAc), 1.54–1.64 (m, 4H, b-CH2, b%-CH2), 1.32–
1.42 (m, 4H, g-CH2, g%-CH2), 1.24–1.32 (m, 6H,
3
3
3
–CH2–); J1,2/1%,2%=10.2, J2,3/2%,3%=9.7, J3,4/3%,4%
=
2.2.1. Sample preparation for lyotropic phase
beha6iour
9.7, 3J4,5/4%,5%=9.7, 3J5,6a/5%,6a%=4.6, 3J5,6b/5%,6b%=2.5,
2J6a,6b/6a%,6b%=12.2 Hz.
The glycolipid samples were prepared as
aqueous dispersions at high buffer content, i.e.
above 80 wt.% using 20 mM Hepes, in some cases
also at lower water content (50%). For this, the
lipids were directly suspended in buffer, and the
suspensions were temperature-cycled several times
between 5 and 70°C and then stored for at least
12 h at 4°C before measurement.
2.1.19. Nonan-1,9-diyl-bis(i-D-thioglucopyra-
noside) 18
1H NMR (400 MHz, d4-MeOH): l=4.27 (d,
2H, H-1, H-1%), 3.78 (dd, 2H, H-6a, H-6a%), 3.58
(dd, 2H, H-6b, H-6b%), 3.15–3.30 (m, 6H, H-3,
H-3%, H-4, H-4%, H-5, H-5%), 3.11 (dd, 2H, H-2,
H-2%), 2.56–2.72 (m, 4H, a-CH2, a%-CH2), 1.56
(mc, 4H, b-CH2, b%-CH2), 1.34 (mc, 4H, g-CH2,
2.2.2. FTIR spectroscopy
3
g%-CH2), 1.22–1.29 (m, 6H, –CH2–); J1,2/1%,2%
=
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 5 min−1. Every 3°C, 50
interferograms were accumulated, apodised,
Fourier-transformed, and converted to ab-
sorbance spectra. The gel to liquid crystalline
3
2
9.7, J5,6b/5%,6b%=5.1, J6a,6b/6a%,6b%=12.2 Hz.
2.2. Methods
Thin-layer chromatography was performed on
silica gel (Merck GF254), and detection was ef-
fected by spraying with a solution of ethanol
sulphuric acid (9:1), followed by heating. Column
chromatography was performed using silica gel 60
(230–240 mesh, Machery and Nagel). Optical ro-

82
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
phase transition LblLa (SB to SA in lamellar
phases) was determined by evaluating the sym-
metric stretching vibration of the methylene
groups ws(CH2) with standard procedures. The
location of ws(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).
In addition, X-ray wide-angle diffraction was
performed in selected cases, in which the determi-
nation of the low-range order is possible. In the
case of a hexagonal array of lipid chains such as
found for many phospholipids, the distance be-
tween neighbouring acyl chains, the lattice con-
stant a, can be calculated from the main
wide-angle reflection at a spacing ratio d by a=2/
ꢀ3×d.
2.2.3. X-ray diffraction
X-ray diffraction measurements were per-
formed at the European Molecular Biology Lab-
oratory (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 with tripalmitin as a standard (periodicity
4.06 nm at room temperature). Further details of
the data acquisition and evaluation system can
be found elsewhere (Boulin et al., 1986). In the
diffraction patterns presented here, the logarithm
of the diffraction intensities log I(s) is plotted
against the scattering vectors. The evaluation of
the X-ray spectra was made according to proce-
dures described 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-dimensional structures. Here lamellar and
cubic structures are of particular relevance,
which can be characterised by the following fea-
tures:
3. Results and discussion
Fig. 4 shows a systematic structure variation
of a classical carbohydrate amphiphile A (dode-
cyl-b-D-glucopyranoside) and depicts clearly the
influence of anomeric linkage, configuration, ring
size and flexibility and electric charges on the
phase stability. Compound A is a monoalkylated
sugar, thus only a smectic A phase can be found.
B (a-gluco), C (a-manno) and D (b-galacto) are
diastereomers of A and the phase behaviour is
very similar, as the configuration seems to have
only a small influence on the thermotropic prop-
erties. The axial substituents increase the clearing
temperatures only slightly, because of the more
rod-like shape of these compounds.
In the glucosamin compound E one OH group
is replaced by a less polar NH2 group, which
acts less efficiently in the hydrogen network.
Thus, the clearing temperature of E is lower rela-
tive to that of A. The fuco derivative F with a
methyl group in the middle of the polar regions
does not show mesophase behaviour, since a
layer structure would not separate the paraffins
from the polar groups. Owing to the higher in-
tramolecular contrast attributable to the car-
boxylic group, the glucoronic acid derivative G
exhibits a higher clearing temperature than A
and the even more polar sulfate group in com-
pound H raises the clearing temperature still
more.
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-dimen-
sional structures. Their various space groups
differ in the ratio of their spacings. The rela-
tion between reciprocal spacing shkl=1/dhkl
The greater flexibility of the five-membered
ring in the glucofuranoside I does not effect the
phase stability: its clearing temperature is nearly
identical with that one of the glucopyranoside A.
Whereas monophilic liquid crystals require stiff
and lattice constant a is shkl=(h2+k2+l2)1/2
/
a (hkl=Miller indices of the respective set of
the plane).

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
83
ring systems, amphiphilic liquid crystals do not
need them.
ent behaviour might be the different possibilities
for hydrogen bonding in the polar headgroup
region, which should be dependent on the
configuration. In this context it is interesting to
compare the clearing temperature of the b-glu-
cosides (compounds B, 4 and 6) with the clearing
temperature of the b-galactosides (compounds D,
14 and 16): If the unpolar part is kept constant,
replacing the glucose by a galactose will stabilise
the SA in all cases by about 20 K.
The introduction of unsaturation in compound
6 decreases the clearing and melting temperature
as expected, already yielding liquid crystallinity at
ambient temperature. If the glucose headgroup is
replaced by a galactose residue, the transition
temperatures are also lowered, but compound 16
is crystalline and not liquid crytalline at ambient
temperature, showing again that a much more
effective network of hydrogen bonding is present
in the galactose compounds. Replacing the stearyl
chain by a cis 9-octadecenyl chain in both cases
(compounds 6 and 16) lowered the clearing tem-
perature by 20 K.
In case of the dodecyl glycerols J and K with
only two OH groups, the intramolecular contrast
is still sufficient for the formation of liquid crys-
talline phases. K has a bulky headgroup and
forms a smetic A phase like the other compounds
while J with an elongated, slim headgroup forms
an ordered smectic B phase, since the diameter of
the headgroup is comparable to the diameter of
the alkyl chain and hence allows a higher degree
of order (Fig. 4).
In the next step we introduced longer alkyl
chains. Table 1 summarises the transition temper-
atures of some long-chain alkyl gluco- and galac-
topyranosides, determined by optical polarizing
microscopy and differential scanning calorimetry
as well as the transition enthalpies.
In the case of the gluco derivatives 2 and 4 the
introduction of six more CH2 groups increases the
clearing temperature slightly with respect to com-
pounds B and A, but effects a slight decrease in
the case of the galacto compound 14 relative to
derivative D. This is in accordance with the gen-
eral concept that the clearing temperature nor-
mally rises with increasing chain length and
reaches a plateau, where an optimal ratio between
the polar and unpolar moieties is achieved, and
drops slightly afterwards. This optimum is
reached in the case of monosaccharides with 14 to
16 CH2 groups and the 12 and 18 chains are
therefore close to this plateau. Since the galacto
compounds with an axial OH group at C-4 are
shorter than the gluco compounds, the optimal
ratio of the polar and unpolar moieties should be
reached somewhat earlier and the compound with
18 CH2 groups is in the case of the galacto
compounds, therefore, further beyond the plateau
than in the case of the gluco compounds.
However, it is remarkable that the gluco and
galacto compounds exhibit the opposite be-
haviour: for the glucosides the a-anomer is the
more stable one, whereas it is the less stable
anomer in the case of the galacto compounds.
There is also a remarkable difference in stability
between the stearyl-a- and b-galactosides 12 and
14, while the stability of the glucosides 2 and 4 is
almost the same. The reason for this quite differ-
The introduction of a stiffer cyclohexyl group
in compounds 8 and 10 increases the clearing
temperatures as should also be expected. It is
somewhat surprising that there is more or less no
difference in stability between the two different
anomers 8 and 10 in this case.
The bola amphiphile 16 did not show any liq-
uid crystalline behaviour, since the linker might
be too short, but it stabilized the liquid crystalline
phase of compounds 2 and 4 in the contact prepa-
ration by 5 and 8.4 K, respectively. The reason,
therefore, might be the increased amount of sugar
in the mixture due to the incorporation of the
bola amphiphile with two sugars and only a short
linking chain, which shifted the ratio between the
polar and unpolar parts in the mixture towards
the sugar. Whereas the optimum ratio and the
maximum clearing temperature is reached in case
of monosaccharide headgroups with 14–16 CH2
groups, it is reached in the case of disaccharide
headgroups with longer alkyl chains. The stearyl
glycosides 2 and 4 are already beyond this opti-
mal ratio because of the excessively long chain.
Upon addition of the bola amphiphile, the
amount of sugar is increased and a better ratio

84
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
Table 1
Transition temperatures and transition enthalpies of alkyl monoglycosidesa
2
Cr 98.0
Cr 97.3 (70.6)
Cr1 93.3
SA 150.8
SA 150.9 (1.2)
SA 146.8
SA 145.8 (0.6)
SA 126.0
SA 181.5
Ib
I
DSC
4
DSC
6
8
DSC
10
DSC
12
DSC
14
DSC
16
Cr2 87.0
Cr2 86.8 (44.0)
Ib
I
Cr3 71.7 (1.7)
Cr1 92.9 (13.8)
Ib
Ib
I
Cr 109.7
Cr3 101.1 (45.0) Cr2 105.8 (7.8)
Cr2 104.5 (4.0)
Cr1 109.0 (20.0) SA 179.7 (0.3)
Cr 124.1 SA 182.5
Cr1 123.7 (24.7) SA 178.9 (2.0)
Cr 89.0
Cr 89.0 (62.6)
Cr 106.0
Cr 105.0 (74.0) SA 160.8 (0.91)
Cr 80.6
Cr 108.9
Ib
I
SA 145.9
SA 144.9 (1.4)
SA 164.2
Ib
I
Ib
I
SA 145.0
Ib
Ic
18
a The DSC data in brackets behind the transition temperature are the transition enthalpies in kJ·mol−1. In some cases additional
transition temperatures Cr (below the crystal to SA transition temperature) and enthalpies are given with the DSC data. They belong
to crystal to crystal transitions, which could not be detected by polarizing microscopy.
b Myelin figures (Bouligand, 1998) in the contact preparation with water.
c Meanwhile, this compound did not show any liquid crystalline phases, it stabilized in the contact preparation the SA phase of
compounds 2 and 4 by 5 and 8.4 K, respectively.

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
85
between the polar and unpolar moieties leads to
an increased clearing temperature.
from the pure thermotropic SA phase in the ab-
sence of water to the lyotropic phase of the fully
hydrated sample in excess water.
3.1. Structural lyotropism
All compounds — visualised in a contact
preparation with water in the polarizing micro-
scope — exhibited only myeline figures, because
of the very low water solubility.
Above the critical micellar concentration gly-
colipids form supramolecular aggregates in
aqueous media (CMC). 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 like pH
and concentration of mono- and divalent cations
(Israelachvili, 1991) Moreover, the molecular
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. Between these
two phase states a reversible transition 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 (mi-
cellar, lamellar L, cubic Q and inverted hexagonal
HII) occurring in glycolipid–water systems is
given by Curatolo (1987), more general informa-
tion about structural polymorphism can be found
elsewhere (Luzzati et al., 1986; Seddon, 1990;
Israelachvili, 1991; Seydel et al. 1993). A compre-
hensive overview of the phases and phase transi-
tions (including the three-dimensional aggregate
structures L, Q, and HII) adopted by glycoglyc-
erolipids is given by Koynova and Caffrey (1994).
The lyotropic behaviour of the compounds pre-
sented in Table 1 was studied by polarizing mi-
croscopy, FTIR spectroscopy and X-ray
diffraction. Whereas the X-ray diffraction mea-
surements were performed in excess water, it is
possible to observe the full range of concentra-
tion-dependent lyotropic behaviour in the contact
preparation with water. When using this tech-
nique a gradient of water concentration is formed
within the sample, showing all phases ranging
3.2. Phase transition of the hydrocarbon chains
In Fig. 5, the results of the acyl chain phase
transition — presented as the peak position of the
symmetric stretching vibration ws(CH2) versus
temperature — are given for the C18-containing
samples directly linked to the sugar backbone.
This compounds melt in the temperature range
55–70°C. Interestingly, the b-glucoside 4 exhibits
higher wavenumber values — higher fluidity or
lower state of order, respectively — at all temper-
atures and also a significantly lower phase transi-
tion temperature Tc than the a anomer 2.
Regarding the latter, this is just the opposite for
the galactose-containing glycolipids (compare the
analogous results above for thermotropic phase
behaviour), for which Tc of the a-anomer 12 is
more than 10°C lower than that of the b-anomer
14, while the fluidity is similar, in particular, in
the gel phase.
The glycolipids with a cyclohexane, but only a
short lipid chain (C8) surprisingly undergo acyl
chain melting at relatively high temperatures (Fig.
6). Furthermore, the a- and b-isomers behave
similarly. The chain melting, however, takes place
over a very broad temperature range (\20°C),
not allowing the determination of a definite Tc. At
the high temperature end the chain melting is still
not completed. This extremely broad range proba-
bly results from the presence of the stiff cyclohex-
ane group directly linked to the flexible alkyl
chain. This group, which is itself hydrophobic,
dramatically increases the hydrophobic moiety
relative to the other substances and is thus re-
sponsible for the high Tc. Concomitantly, the
broadness of the transition may be understand-
able. A favourable molecular packing of the hy-
drophobic part, necessary for cooperative melting,
is impeded drastically because of the voluminous
cyclohexane group (Fig. 6).

86
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
Fig. 5. Phase transition of the hydrocarbon chains of glucose- and galactose-containing glycolipids bound to a C18 chain in a- or
b-linkage. The peak position of the symmetric stretching vibration of the methylene groups lies in the range 2848–2850 cm−1 in gel
or crystalline phases and in the range 2852–2854 cm−1 in liquid crystalline states.
Fig. 6. Phase transition of the hydrocarbon chains of glucose-containing glycolipids 8 and 10 bound in a- and b-linkage to a
cyclohexan-C8 chain (see also legend of Fig. 5).

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
87
Fig. 7. Wide-angle X-ray scattering (WAXS) patterns of gal-a-OC18 in 90% buffer at selected temperatures.
Fig. 8. Small-angle X-ray scattering (SAXS) patterns of glu-a-OC18 and glu-b-OC18 in 90% buffer at selected temperatures.
To determine the packing density of the acyl
chains and to verify or disprove the above hy-
potheses, X-ray wide-angle diffraction measure-
ments were also performed using gal-a-OC18
(stearyl-a-galactose, Fig. 7). Between 5 and 60°C,
sharp reflections are observed at 0.367–0.370,
0.452, and 0.248 nm. From the appearance of these
peaks (in the Lb phase, usually only one reflection
at 0.415 nm is observed), the existence of a ‘crys-
talline’ Lc phase found, for example, for diacylglu-
coses can be concluded. At 80°C, the peaks at the
lower spacing ratios have vanished, and the re-
maining weak and broad reflection at 0.455 nm is
typical for the occurrence of the a-phase.
Also, the cyclohexane glucosides exhibit various
sharp diffraction maxima typical for the occur-
rence of the Lc phase. In particular, the a-linked
anomer has maxima at 0.333, 0.357, 0.426, 0.451,
and 0.486 nm at 5°C. These reflections are basi-
cally present — with slight shifts of the peak
position — up to 60°C, and vanish above this
temperature.

88
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
3.3. Supramolecular aggregate structures
with the respective patterns obtained at 50%
buffer content. At 90% buffer content and 40°C
(Fig. 9A), two periodicities at 3.53 nm and 3.11
nm and the corresponding second order reflec-
tions are clearly observed. This is similarly true
for the sample at 50% buffer content and 40°C
(periodicities at 3.51 and 3.07 nm, Fig. 9B and
Table 2). However, this pattern contains further
reflections, which may indicate another substruc-
ture. For example, the reflection at 1.43 nm=3.51
nm/ꢀ6 might be indicative of a cubic substruc-
ture. This interpretation seems to be improbable
because it is usually believed that cubic phases
occur exclusively in the fluid phase of the hydro-
Fig. 8 shows X-ray diffraction patterns for
stearyl-a-glucoside 2 and stearyl-b-glucoside 4 in
90% buffer and at different temperatures.
Both samples exhibit similar patterns with
reflections at equidistant ratios in the temperature
range 20–60°C, which convert at 80°C into dif-
fraction patterns with reflections lying again in
equidistant spacing ratios, but with a much higher
periodicity. For a more exact characterization, in
Fig. 9 the pattern of stearyl-a-glucoside at 90%
buffer content at the temperatures 40 and 80°C,
i.e. below and above Tc are presented together
Fig. 9. SAXS patterns of glu-a-OC18 at two buffer contents (50 and 90%) and two temperatures (40 and 80°C).
Table 2
Values for the measured periodicities of the stearyl-monosaccharides in nm
Stearyl-a-glucoside
Stearyl-b-glucoside
Stearyl-a-galactoside
Stearyl-b-galactoside
3.52, 3.11
3.26
3.45
4.95
4.46
4.49
4.58
3.51, 3.07
3.26
3.44
4.90
4.00, 3.26
4.46
3.51
3.51
4.49, 4.24

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
89
Fig. 10. SAXS patterns of glu-a- and glu-b-cyclohexane compounds 8 and 10 in 90% buffer at selected temperatures.
carbon chains (Luzzati et al., 1986). However, for
particular glycolipids, bacterial lipid A and lipo-
polysaccharide, the existence of cubic phases
could be proven already in the gel phase of the
acyl chains (Brandenburg et al., 1990, 1998). Of
course, because of the lack of further reflections
indexing on such cubic structures, at the moment
no nearer characterisation of this phase is
possible.
The periodicities at 80°C (4.90–4.95 nm) may
be assumed to be typical for the occurrence of a
‘normal’ L multilamellar phase, whereas the val-
ues below 80°C indicate an interdigitated lamellar
phase. The occurrence of interdigitated phases is
observed for diacyl phospholipids only at high
ionic concentration or by adding polar sub-
stances, as found by Boggs and Rangaraj (1985).
However, for particular glycolipids, i.e. lipid A,
such interdigitation has been observed also for the
pure lipid in the gel phase (Brandenburg et al.,
1990). In the case of the investigated monoalkyl
glycolipids, the reason for the occurrence of such
an interdigitated phase is the disproportion be-
tween the cross-section of the polar and apolar
moieties allowing two packing organisations: as
micelles with the apolar chains packed together in
the interior of a sphere and the polar headgroup
lying on the surface, or the packing in a lamellar
arrangement with interdigitated chains, which
now resembles the situation of a ‘normal’ diacyl
phospholipid. In both cases, a minimisation of the
hydrophobic free energy is guaranteed. Although
for monoacyl glycolipids interdigitated phases
have not been described up to now, the existence
of such a phase has been proven for 1-stearoyl-
lysophosphatidylcholine (Mattai and Shipley,
1986), a compound resembling the investigated
glycolipids with respect to the disproportion of
polar and apolar moieties. In accordance with the
occurrence of such interdigitation is the known
tendency of lyso phospholipids to incorporate
spontaneously into target cell membranes thus
again minimising the Gibbs free energy.
In the liquid crystalline phase the tight packing
is lost due to the strong occurrence of gauche
conformers leading eventually to a ‘normal’ La
phase with the terminal methyl groups facing each
other. This general behaviour was found to be

90
V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
Bouligand, Y., 1998. Defects and textures. In: Demus, D.,
Goodby, J., Gray, G.W., Spiess, H.-W., Vill, V. (Eds.),
HandbookofLiquidCrystals, vol. 1. Wiley-VCH, Weinheim,
pp. 406–453.
Boullanger, P., 1998. Amphiphilic carbohydrates as a tool for
molecular recognition in organized systems. Top. Curr.
Chem. 187, 275–312.
Boulin, C., Kempf, R., Koch, M.H.J., McLaughlin, S.M., 1986.
Data appraisal, evaluation and display for synchrotron
radiation experiments: hardware and software. Nucl. Instr.
Methods A249, 399–407.
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 behaviour
of phospholipid membranes studied by infrared spec-
troscopy. Biochim. Biophys. Acta 779, 381–401.
Curatolo, W., 1987. The physical properties of glycolipids.
Biochim. Biophys. Acta 906, 111–136.
Ellens, H., Siegel, D.P., Alford, D., et al., 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.),
HandbookofLiquidCrystals, vol. 3. Wiley-VCH, Weinheim,
pp. 341–392.
true for all other stearyl monosaccharides, and the
values for the measured periodicities are listed in
Table 2.
The reason for the occurrence of two sets of
lamellar reflections for stearoyl-¦-glucose (Fig. 9) is
not completely clear. We can exclude an inhomo-
geneity of the chemical structure, and also inhomo-
geneoushydration, thelatterbecauseoftherepeated
heating/cooling cycles prior to measurement guar-
anteeing complete hydration. The observation that
the two sets of lamellar reflections are invariant in
the temperature range from 20 to 60°C, however,
also speaks against this hypothesis. Interestingly,
this effect is very specific for stearoyl-a-glucose, and
is not found for any other compound investigated.
The two cyclohexane-containing glycolipids as-
sume multilamellar structures under all conditions
investigated (Fig. 10A and B). For the b-linked
compound the change of the lamellar periodicity
from 3.69 to 4.39 or 4.50 nm, respectively, between
40 and 60°C (Fig. 10B) apparently indicates a
transition from an interdigitated to a ‘normal’
lamellar phase. This transition is not observed for
the a-linked compound (Fig. 10A), although both
isomers have a similar state of order in the inves-
tigated temperature range (Fig. 6).
8
Fischer, E., Helferich, B., 1911. Uberneuesynthetischeglycoside.
Liebigs Ann. Chem. 383, 68–91.
Gabriel, A., Dauvergne, F., 1982. The localization method used
at EMBL. Nucl. Instr. Methods 201, 223–224.
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.
Acknowledgements
This work was supported by a grant of the DFG
(SFB470). We are also grateful to the Studiens-
tiftung des Deutschen Volkes for a grant.
Koch, R., 1884. Die aetiologie der tuberkulose. Mitt. K.
Gesundh. 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 biological
relevance. Biochim. Biophys. Acta 988, 221–256.
Luzzati, V., Gulik, A., Gulik-Krzywicki, T., Tardieu, A., 1986.
Lipid polymorphism revisited: structural aspects and biolog-
ical implications. In: Lipids and Membranes: Past, Present,
and Future. Elsevier, Amsterdam, pp. 137–151.
Mattai, J., Shipley, G.G., 1986. The kinetics of formation and
structure of the low-temperature phase of 1-stearoyl-
lysophosphatidylcholine. Biochim. Biophys. Acta 859, 257–
263.
References
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.
Bo¨cker, T., Lindhorst, T.K., Thiem, J., Vill, V., 1992. Carbohydr.
Res. 230, 245–256.
Boggs, J.M., Rangaraj, G., 1985. Phase transitions and fatty acid
spin label behavior in interdigitated lipid phases induced by
glycerol and polymyxin. Biochim. Biophys. Acta 816, 221–
233.
Prade, H., Miethchen, R., Vill, V., 1995. Thermotrop flu¨s-
sigkristalline kohlenhydrat-amphiphile. J. Prakt. Chem. 337,
427–440.

V. Vill et al. / Chemistry and Physics of Lipids 104 (2000) 75–91
91
Seddon, J.M., 1990. Structure of the inverted hexagonal (HII)
phase, and non-lamellar phase transitions of lipids.
Biochim. Biophys. Acta 1031, 1–69.
Seydel, U., Labischinski, H., Kastowsky, M., Brandenburg,
K., 1993. Phase behaviour, supramolecular structure, and
molecular conformation of lipopolysaccharide. Immuno-
biol. 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.
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., et al., 1989. Physiolog-
ical levels of diacylglycerols in phospholipid membranes
induce membrane fusion and stabilize inverted phases.
Biochemistry 28, 3703–3709.
Stanger, P., Vill, V., Rohde, S., Jeschke, U., Thiem, J., 1994.
Mesogenic amino sugars. Liq. Cryst. 17, 589–595.
Tietze, L.F., Bo¨ge, K., Vill, V., 1994. Liquid-crystalline
D-glu-
Siegel, D.P., 1986b. Inverted micellar intermediates and the
transition between lamellar, cubic and inverted hexagonal
lipid phases. II Implications for membrane-membrane in-
teractions and membrane-fusion. Biophys. J. 49, 1171–
1183.
cose dialkyl acetals and dodecyl
Chem. Ber. 127, 1065–1068.
Vill, V., Bo¨cker, T., Thiem, J., Fischer, F., 1986. Studies on
liquid crystalline glycosides. Liq. Cryst. 6, 349–356.
Vill, V., Lindhorst, T.K., Thiem, J., 1991. Liquid crystalline
6-desoxy glycosides. J. Carbohydr. Chem. 10, 71–786.
D-glucofuranosides.
Siegel, D.P, 1986c. Inverted micellar intermediates and the
.
.