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Liquid crystalline phases given by helical biological polymers (DNA, PBLG and xanthan). Columnar
textures
F. Livolant, Y. Bouligand
To cite this version:
F. Livolant, Y. Bouligand. Liquid crystalline phases given by helical biological polymers (DNA, PBLG and xanthan). Columnar textures. Journal de Physique, 1986, 47 (10), pp.1813-1827.
�10.1051/jphys:0198600470100181300�. �jpa-00210378�
Liquid crystalline phases given by helical biological polymers (DNA, PBLG
and xanthan). Columnar textures
F. Livolant and Y. Bouligand
CNRS and EPHE, 67 rue Maurice-Günsbourg, 94200 Ivry-sur-Seine, France (Reçu le 27 novembre 1985, accept6 sous forme définitive le 17 juin 1986)
Résumé. 2014 Des phases cristallines liquides hexagonales ont été obtenues avec des polymères hélicoïdaux d’intérêt biologique, essentiellement l’ADN et le PBLG, en solution très concentrée. La plupart des textures classiquement décrites dans les phases hexagonales ont été également observées avec ces polymères (textures
en éventail, textures riches en disinclinaisons). D’autres sont nouvelles, comme les textures à ondulations ou
les empilements hélicoïdaux de domaines hexagonaux. Ces textures nouvelles semblent liées à la nature même des molécules (longueur et forme en hélice). Cependant, l’organisation hélicoïdale et l’ordre hexagonal sont incompatibles et cette question est discutée : ces organisations permettraient de relaxer de manière continue
ou discrète le twist qui se manifeste spontanément entre molécules hélicoïdales et qui est empêché par l’ordre
hexagonal. Enfin, la nucléation de la phase hexagonale au sein de la phase cholestérique et sa croissance ont été suivies au microscope polarisant. Le c0153ur des lignes de défaut présentes dans la phase cholestérique jouerait le rôle de centres de nucléation et les textures hexagonales dépendraient des textures cholestériques
au sein desquelles elles ont grandi.
Abstract. - Highly concentrated solutions of helical polymers of biological interest, namely DNA and PBLG, have been found to form hexagonal liquid crystalline phases. Most of the classic textures of hexagonal phases
are shown by these polymers (fan-shaped textures, textures with numerous disclinations). In addition there are
other new textures such as the undulating patterns and the helical stacking of hexagonal domains. These new textures are probably a consequence of the helical nature of the molecules. However, perfect extended
helicoidal organization and hexagonal order are incompatible. We shall discuss the way in which these new textures allow a relaxation of the twist which occurs spontaneously between helical molecules (and which is incompatible with extended hexagonal order) in a continuous or abrupt manner. Finally, the nucleation and
growth of the hexagonal phase within the cholesteric phase were observed in the polarizing microscope. The
cores of the defect lines which exist in the cholesteric phase act as nucleation centres and the hexagonal
textures formed depend on the nature of the cholesteric textures surrounding them.
Classification
Physics Abstracts
61.30 - 61.70
1. Introduction.
Numerous polymers are known to form liquid crys-
talline phases. Among them are polymers of biologi-
cal interest, which represent a significant part of biological tissues. The study of their liquid crystalline properties and textures is necessary for an understan-
ding of their behaviour in living systems and this topic has not been extensively investigated in the
past.
With this aim in mind, we selected for study the liquid crystalline phases of the three biopolymers :
DNA (a polynucleotide), PBLG (a polypeptide) and
xanthan (a polysaccharide).
These polymers form lyotropic liquid crystalline phases whose structures depend on the polymer
concentration. When the concentration is increased,
cholesteric spherulites appear within the isotropic phase. The appearance of PBLG spherulites was
described by Robinson et al. [1] and explained by
Pryce and Frank (in the appendix of 1). Subsequently
numerous compounds (biopolymers and small liquid crystal molecules) were found to give similar spheru-
lites (Bouligand and Livolant [2]). At higher concen- trations, the cholesteric regions expand and ultima-
tely occupy the whole preparation. The textures and
defects of this phase have been analysed [3]. When
the polymer concentration increases further, the cholesteric organization disappears and is replaced by another liquid crystalline phase whose textures
and defects are described here. Analysis reveals that this phase is columnar and probably hexagonal.
These observations are supported by X-ray diffrac-
tion data which show that highly concentrated prepa- rations of DNA and PBLG have local hexagonal
order [1, 4-8]. The textures of this hexagonal liquid crystalline phase have not been previously analysed.
They are described here and compared with those
exhibited by classical (i. e. amphiphilic) lyotropic phases. Most textures were similar but some ones
Article published online by EDP Sciences and available at http://dx.doi.org/10.1051/jphys:0198600470100181300
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new arose with an undulating appearance. These
new textures are related to the problem of the
coexistence of hexagonal and helical ordering.
The transition between the cholesteric and the
hexagonal phase was studied optically and hexagonal
textures were found to depend on cholesteric textu- res within which they had nucleated.
2. Material and methods.
2.1 DNA. - Calf thymus DNA (Merck) was solubi-
lized in 15 mM Tris-Cl- buffer, pH 8 and sonicated
to obtain short fragments whose length was less than
3 )JLm. The methods used to obtain liquid crystalline phases have been described in a previous work [3].
A cholesteric organization appeared first and was
replaced by the hexagonal phase several hours or
days later.
2.2 PBLG. - Two kinds of Poly-ybenzyl-L-gluta-
mate were used : PBLG MM 20 000, synthesized by
the Centre of Molecular Biophysics (Orldans, France) and PBLG MM 28 000 (type III, Sigma).
They were dissolved in dioxan.
2.3 XANTHAN. - This polymer secreted by Xantho-
monas campestris was purified and kindly provided by Drs. Rinaudo and Milas (CERMAV, CNRS, Grenoble, France). This polysaccharide (MM 200 000) was used in aqueous solutions (distilled
water, 6 to 60 % NaCI or KCI solutions).
All preparations were observed with a polarizing microscope (Orthoplan Pol Leitz and Optiphot X
Pol Nikon) either between crossed polars or
between opposite circular polarizer and analyser.
3. Hexagonal phases in liquid crystals (reminders).
Hexagonal liquid crystalline phases are classically
formed by amphiphilic compounds, such as phospho- lipids. In an appropriate range of temperature and concentration, molecules form columns which align
in parallel and often show a hexagonal order in
section (Fig. la, b). Numerous works deal with the structure and phase diagrams of these lyotropic liquid crystals [9, 10].
More recently discoid molecules made of a rigid
core surrounded by flexible paraffinic chains have
been synthesized. These molecules tend to stack in
more or less ordered columns which are arranged in
a hexagonal array (Fig. lc). Each column is able to slide longitudinally with respet to the others. These
phases are called hexagonal discotics ; their textures
have been much studied [11-14]. Hexagonal ordering
has also been found within smectic layers. The hexagonal order is often coherent over several layers
and in this case, the phase can be considered to be true crystal (Fig. Id).
Textures exhibited by DNA and PBLG strongly
resemble the classical textures presented both by amphiphilic and discotic molecules. In this case, the
elongated molecules are aligned in parallel and form
a hexagonal array. Each molecule is able to translate
Fig. 1. - Various hexagonal liquid crystalline phases
formed by very different kinds of molecules. a, b) lyotro- pic hexagonal phases of amphibilic molecules. a) direct hexagonal phase, b) inverse hexagonal phase. c) discotic hexagonal phase : disc-shaped molecules are stacked and form columns which are arranged in a hexagonal array.
d) smectic B phase : molecules are elongated rods orien- ted normally to the layers and forming a hexagonal
network. e) polymeric molecules (usually helices) are aligned in parallel with the ability to move longitudinally.
f) transverse section of a hexagonal array of elongated
molecules. There are numerous axes of symmetry: two- fold axes (L2, T2 and a2), three-fold axes (L3) and six-fold
axes (L6).
longitudinally and to rotate about its long axis (Fig. 1 e).
Symmetries of hexagonal columnar liquids are
reviewed in figure If. Most symmetry axes lie parallel
to the molecular axes : six-folds axes L6 , three-
fold axes ( L3 ) and two-fold axes L2 . The other
two-fold axes are normal to molecules (T2 and 6z).
Actually, the presence of a L6 axis has been proved
from the analysis of textures given by certain discotic
compounds [12]. A L6 axis probably exists also in the
phases of DNA and PBLG studied here. For symme- tric molecules, each point of L2 or L6 is a centre of symmetry and each plane normal to molecules is a
plane of symmetry as well as L6, T2 and Lb, 02 . This cannot be true for hexagonal phases
given by asymmetric molecules such as helical poly-
mers.
4. Results.
4.1 HEXAGONAL COLUMNAR TEXTURES. - Nume-
rous textures presented by PBLG (Pl. I) and DNA (Pl. IIa-d) are characteristic of hexagonal liquid crystalline phases. Others are new and perhaps specific to long helical polymers. All these textures
are more difficult to analyse in DNA since the domains are smaller and therefore the details within them do not appear so precisely. However, they
appear highly similar in the two polymers and will be considered together.
Undulating textures were observed with the two
polymers and two situations may be considered :
Plate I. - Hexagonal textures of PBLG. a-c : Transformation from undulating patterns (a) to herring-bone patterns (c)
when the polymer concentration is increased. a : Undulating patterns showing numerous double defects. Dark lines follow loci of maximum curvature of molecular orientations (x 120 ; + ). b : Loci of maximum curvature of molecules transform into walls of discontinuity and their path becomes sinuous (arrows). New undulations appear in each
elongated domain (x 180 ; + ). c : Herring-bone pattern (x 300 ; + ). d : Other undulating patterns. The different domains are interrupted by walls (w). Illuminated regions draw either elongated rectangles (in the central domain) or elongated triangles (on the right) (x 300 ; 4+- ). e : Hexagonal patterns observed in free-surface drop. Molecular
orientations are underlined by fine striae (s). Certain walls showing stairs patterns are clearly observed (w) (x 300 ; + ). f, g : Twist effect observed between hexagonal domains in highly concentrated preparations. f : Beginning of a double spiral is created by the helical stacking of the domains in free-surface drop (x 300 ; + , 2 ’B/4). g : Series of arcs produced by the same process, between slide and coverslip (x 330 ; + ).
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Plate II. - Hexagonal textures of DNA obtained in presence of KCI (a, d) or in «/1 conditions (b, c). a : Fan-shaped
textures (x 450 ; + ). b : Domains showing striations radiating around numerous centres (x 925 ; + ). c : Undulating patterns (x 1725 ; + ). d : Other undulating patterns interrupted by walls (w). Other walls (w’) are underlined by a
series of dark triangles (x 1505 ; + ). e : Liquid crystalline phase, probably hexagonal, obtained with xanthan in aqueous solution (x 390 ; +).
- Undulations may occur in the plane of the preparation, i.e. around disclination lines normal to this plane. Such textures appear therefore as an alternation of light and dark elongated domains
which transform into bright regions interrupted by
thin dark parallel lines when the microscope stage is rotated (Pl. la, b ; Pl. IIc). Molecular orientations in such domains are indicated in figure 2. In (c) the sine
curves representing the distribution of DNA molecu- les extend parallel to one polar and extinctions are
observed only at the minima and maxima of the undulations. The dark bands extend and move when the microscope stage is slightly rotated in one
direction or in the other (b, d) and transform into
larger dark bands whose number is reduced by a
factor of 2 and which alternate with light bands (a, e).
Defects occurring in such undulating patterns are always double (Pl. la) but among the three possible
situations schematically represented in figure 3, only
the first and the third ones are observed, as revealed
Fig. 2. - Undulating patterns observed between crossed
polars (P, A). In (c) all of the domain appears bright except the thin lines along where molecules are aligned parallel to one of the polar directions. When the micros- cope stage is rotated (in either direction), the dark lines
extend and move (b, d) and transform into an alternation of light and dark large bands (a, e).
Fig. 3. - Defects occurring in helical systems. Two layers (or one period P) are added simultaneously either at
a point of maximum curvature (a, b) or at an inflexion point (c), giving either branched patterns (a, b) or flame patterns (c) when viewed between crossed polars. In the vicinity of the core of defects, the distortions of the director are slightly exagerated to make clear the drawing.
by the use of a quartz first order retardation plate.
When the polymer concentration is increased the dark lines following the loci of maximum curvature of molecules transform into walls of discontinuity
and their path becomes sinuous (Pl. Ib). In each elongated domain a series of parallel flames appear
obliquely with respect to the elongation axis of the domain (Pl. Ia). This reveals new molecular undula- tions whose orientation is more or less normal to the flame axes. These undulations create new walls, parallel to the flame direction, which divide each domain into numerous microdomains. This develop-
ment, schematically represented in figure 4, creates herring-bone patterns such as those presented in plate Ic. Numerous more complicated textures obser-
ved in PBLG are also formed by this iterative process, repeated three times or more, recalling
certain fractal structures.
- The undulating patterns are quite different
when the + zr disclination lines (6) lie more or less parallel to the preparation plane : large regions
limited by walls (w) present a transverse striation
corresponding to progressive variations of the illumi- nation (Pl. Id, IId). In such regions, the defect lines
are more often virtual, alternatively above and
below the preparation plane which creates the
undulation effect (Fig. 5a). Two simple illustrations of this organization are given in figure 5b-e. In (b) 8
is horizontal and bands of equal illumination corres-
ponding to similar orientations of molecules form
elongated rectangles which are arranged symmetri- cally on both sides of the defect line (c). In (d) 6 is oblique and the two bands present a trapezoidal shape and converge (e).
The walls which cross these undulations patterns
are probably twist walls. In figure 6 they are repre- sented in top view (a), as observed in the microscope
and in vertical section along the wall (b). Columns
are shown in front of and behind the wall and the
corresponding twist is either positive or negative. In
these figures, 5 lines are virtual. When the 5 lines are
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Fig. 4. - Transformation of undulating patterns (a) into herring-bone patterns (d). Thin continuous lines represent the molecular orientations. Walls of bend are indicated by
dotted lines (L) and walls of discontinuity are symbolized by full dark lines (W). The walls of bend (L1) observed in the undulating patterns and revealed in the microscope by
dark lines moving when the stage is rotated (a) transform
into walls of discontinuity (W1) when the concentration is increased (b). The same process occurs again in each elongated region by the formation of new undulations revealed by new walls of bend (L2) (c) which transform
again into walls of discontinuity (W2) (d).
present in the liquid crystal, the twist is stronger and this corresponds to the situation of figure 7 : the perfect 6 line represented in (a) which is an edge-
disclination + or is broken in (b) and a twist wall is
created. Theoretically, the two separated half-lines
are joined by a twist disclinaltion, but a twist wall is
more likely. Indeed, a micrograph published in [14]
illustrates this effect : the wall extends largely on
both sides of the disclination and progressively
vanishes at its extremities, this probably corresponds
to a decrease of the twist, as shown in lateral parts of figure 7c.
Fig. 5. - a) Schematic representation of textures undu- lating between planes of slide and coverslip around
numerous + 7T disclinations. We have only drawn the planes along which molecules are lying. The defect line 5 is
parallel (b, c) or slightly oblique (d, e) with respect to the observation plane. This line may lie in the preparation plane or be virtual which is represented here. In the first
situation (b), molecular orientations are identical at a
given distance of 5 ; bands of equal birefringence have a
defined width and lie parallel to 6 (c). When 8 is oblique (d), these bands are convergent and of variable width (e).
Fig. 6. - a) An apical view of a region with undulations
(such as those schematically shown in Fig. 5a). Numerous
+ ir disclinations are created around lines (8) which are oblique with respect to the preparation plane. Moreover
these domains are interrupted by twist walls (W) such as
those described in figure 7. b) Side view of a twist wall
(W) presented in a. The twist is alternatively positive and negative between molecules of the two domains (dotted
lines and continuous lines). Second order walls (X’ and X") correspond to inversions of the curvature radius of molecu- lar orientations and cannot be observed in the micro-
graphs.
Fig. 7. - The line of a + ir disclination (8) is interrupted by a cleavage surface S normal to this line (a). A lateral displacement of the two parts of the fault (b) creates a
twist wall. An apical view of this region (c) reveals that the twist angle is important in the region separating the two
disclination lines (8 and 8’) whereas it is much smaller on
both sides of it.
Some regions recall the broken fan-shaped textures
of smectic phases and are obtained either between slide and coverslip (Pl. IIa, b) or in free surface drops (Pl. Ie). Each fan is made of a series of narrow
sectors, differently contrasted in the polarizing microscope, and converging at a common centre. In
free surface drops, molecular orientations are easily
followed since they are underlined by a fine striation which runs normally to the radial sector boundaries
(Pl. Ie). These fans may be nearly closed (Pl. IIa) or slightly open (Pl. Ie, IIb) and they are abruptly interrupted by walls which present stairs patterns
(Pl. Ie). These walls separate two striated domains radiating around two centres Cl and C2. The direc-
tion of the wall (w) crosses the line joining Cl and C2 at an angle of 10 to 15° (Fig. 8a). Along w,
molecular orientations do not vary significantly in D2 (a 2 remains not far from 90°) but changes in D1.
Stairs walls are observed when a 1 is between 60 and 85°. (For highest values of « 1 there is no wall since
« 1 + a 2 is close to 180° and for smallest values, these
walls transform into walls of developable domains [14]). The distribution of molecular orientations is
presented in figure 8b. The wall is divided into small
oblique segments which are walls of discontinuity on
both parts of which molecular orientations are
symmetric. Each step corresponds to a discrete
number of added columns and two possible distribu-
tions of columns may be proposed in these regions
which form the complementary series of segments (Fig. 8c, d). The situation c is the more probable
since a symmetric distribution of molecular orienta- tions would appear to minimize the energy of the wall.
Another kind of wall, revealed by differences of contrast in the polarizing microscope, presents a radial distribution in each domain (Pl. Ie). Such
walls correspond to changes in the molecular orienta- tions and they are reminiscent of the patterns described by Merrucci [15] for hexagonal discotics.
Hexagonal textures showing numerous disclinations
were obtained in PBLG (Pl. IIIe, f). The planar
Fig. 8. - a) A stair wall separates two striated domains
Di and D2 in which molecular orientations radiate around two centres Cl and C2. The wall is located between the two
arrowheads, in the region where molecular orientations in the two domains make an angle ranging from 150 to 185°.
b) Molecular distribution along this wall which is divided into two series of segments. The first series corresponds to
walls of discontinuity on both sides of which molecular orientations are symmetric. These segments are therefore oblique with respect to the wall direction (W). c, d) Two
situations may be proposed for the second series of segments, the first one (c) being more probable since it coresponds to a symmetric distribution of molecular orientations.
texture of the hexagonal phase grows in the homeo-
tropic solution around vertical + 7T disclinations.
Molecular orientations rotate by about 180° around
each of them and walls are very often (but not invariably) linked to these defect lines. In these
disclinations, the defect line (6) lies locally parallel
to the rotation axis (f2) and these two directions are normal to molecular orientations (n). In this case,
the rotation axis follows either the T2 or 62 axis of
the hexagonal network. The rotation may occur without formation of a wall (Fig. 9a, b) but this
situation is rarely observed in PBLG and DNA.
Indeed, walls are frequently anchored to the defect
line and they usually lie in the plane of symmetry of the disclination. Several possible situations may be considered :
(i) A vertical displacement is formed after the 180° rotation and creates a twist boundary which can
relax into a series of screw dislocations (Fig. 9c, d).
(ii) The rotation is less than 180° (Fig. 9e) which produces grain boundaries in planes both parallel
and normal to the molecules.
(iii) The rotation axis (f2) and defect line (6) are
different from T2 or e2, with 111 parallel to & This also produces grain boundaries in vertical and hori- zontal planes (Fig. 9f). From the appearance of the
