Food Science: Professor Chen Shan, Guangxi University, et al.: Research progress on self-assembly of gel polysaccharides and their applications

As a biological macromolecule composed of basic monosaccharides, polysaccharides have the diversity of linkages, straight/branched chain ratios and molecular masses, as well as molecular polyhydroxylones and some special spatial structures, which provide good conditions for
their functionalization.
Self-assembly is the process of macromolecules to drive molecules or parts of molecules to spontaneously form ordered structures through non-covalent bond interactions, and it is also the basis for the formation of
supramolecular structures through interaction between biological macromolecules.
Compared to proteins, polysaccharides are more stable under a variety of environmental factors (temperature, pH, etc.
) and can be combined with other functional factors to construct functional complexes
with ordered structural and specific advantages.

As a triple helix polysaccharide, gel polysaccharide (CUR) is a linear β-(1,3)-D-glucan, which was first obtained by fermentation of alkaline-producing bacillus faeces, and was approved by the US Food and Drug Administration as a food additive, with excellent thickening and water retention making it widely used
in the field of food processing.
Pan Yuxue, Xu Xindong, Chen Shan* from the College of Light Industry and Food Engineering, Guangxi University, reviewed the characteristics of CUR's self-assembly and the function and biological activity of constructing functional materials based on self-assembly, which provided a reference
for the in-depth research and application of CUR's self-assembly in food and other fields.

1.
CUR triple helix conformation

As shown in Figure 1A, the CUR has a 6/1 right-handed helical configuration, the main chain is connected by β-1,3-bonds, and every 6 glucose residues form a spiral, and the intramolecular hydrogen bond between C4 and C6 hydroxyl groups on the backbone glucose ring is the driving force
for the formation of the helical structure 。 As shown in Figure 1B, the triangular hydrogen bond network located in the same xy plane and perpendicular to the helical axis is formed by the intermolecular hydrogen bonding between the hydroxyl groups on C2, and the three polysaccharide linear chains form a tight cross-helical structure under the action of intermolecular and intramolecular hydrogen bonds, and the helical center is considered to have an unequal sided "hexagonal" hydrogen bonding in the current mainstream view, that is, 6 atoms of 3 hydroxyl groups on the 3 sugar chains form an unequal hexagonal hydrogen bond model to form a stable COR triple helix structure
。 It has been reported that the C2 hydroxyl group on the main chain of Schistomycetes, which also has a triple helix conformation, is hydrophobic, while the C6 hydroxyl group and side chain glucose groups are hydrophilic, which makes the hydrophobic surface always located in the triple helix structure when the single chain is restored to the triple helix conformation, and the formed triple helix structure is covered by a hydrophilic surface, forming a hydrophobic cavity with a diameter of about 3.
5 Å within the triple helix structure, and the denatured cavity is flexible
。 For CUR, the center of its helical structure is occupied by the C2 hydroxyl group, resulting in the inability of a hydrophobic cavity to form; In addition, the presence and number of glucose-based side chains can not only significantly change the physical properties of dextran, but also have a strong influence on the hydrophobic cavity of the triple helix structure, the higher the degree of branching, the shorter the pitch formed, and the larger the volume of the
hydrophobic cavity.
Therefore, compared to other triple helix polysaccharides with side chains, the triple helix conformation formed by the self-assembly of the CUR does not have a hydrophobic cavity
.
In addition to the Type A hydrogen bonding model, different hydrogen bonding models have been proposed in the predecessors, in which in the Type B hydrogen bonding model, it is believed that the CUR sugar chain is right-handed, and an intramolecular hydrogen bond is formed between the C2 hydroxyl groups of adjacent sugar rings on the same chain (Figure 1C), and the driving force for the formation of the triple helix structure may be van der Waals force; In the Type C hydrogen bonding model, unlike the other two models, the CUR triple helix is intermolecularly left-handed, and the hydrogen bond network formed by the intermolecular hydrogen bond between C2 hydroxyl groups is not perpendicular to the helix axis, but follows the helix (Figure 1D).

Among these three models, the most unstable is the Type B hydrogen bonding model, while the Type A hydrogen bonding model is considered to be a more reasonable triple helix conformational model
.

2.
Self-assembly of gel polysaccharides under different induction methods

As shown in Figure 2, the polysaccharide triple helix can be unfolded in the form of random coiling in strongly polar solvents such as dimethyl sulfoxide (DMSO), a process called the unwinding of the triple helix, when the polarity of the solution is reduced, random coiling can re-form the triple helix through specific hydrogen bond recognition, a process called the triple helix rerotation
.
CUR initiates triple helix unrotation under different induction conditions, and by changing the conditions of the solution, the random coil formed after dismantling can be re-formed by re-spinning
.
However, the self-assembly process exhibited by CUR under different induction conditions was slightly different
.

Self-assembly of CUR under heating/annealing treatment

CUR, also known as hot gel, has unique gelling properties that when heated above 80 °C and annealed, a thermoirreversible gel with high gel strength will be formed, and when heated to 55~65 °C and annealed, a thermoreversible gel with low gel strength will be formed (Figure 3).

。 At present, the mainstream view is that the gel mechanism of the former is microscopicly manifested as the triple helix structure is derotated into a single chain at high temperature, and the triple helix structure is re-rotated after annealing to form a triple helix structure, and the three helix structures are connected by at least one single strand or exist in the form of aggregates, which has a strong network structure, thus showing extremely high gel strength at the macroscopic level; The gel mechanism of the latter is microscopicly manifested as a triple helix that is unrotated into a single helix under heating, and a small number of single helix respin is a triple helix during annealing, but the solution is still dominated by a single helix, and there is also a certain connection between multiple single helix or triple helix chains, but the strength of the network structure formed is not as good as the former, thus showing a lower gel strength
at the macroscopic level.

Self-assembly of COR in dimethyl sulfoxide/aqueous solution system

It can be seen from Figure 4 that when the DMSO mass fraction in the DMSO/water system is high, the three heliaxes of the related polysaccharides will undergo conformational transformation, and all exist in
the solution in random coiling form after the DMSO mass fraction reaches 100%.
It is worth noting that with the increase of the molecular mass of the triple helix polysaccharides, the mass fraction of DMSO with different triple helix polysaccharides to maintain the triple helix conformation also increases, which indicates that in the DMSO/aqueous solvent system, when the molecular mass increases, the number of hydrogen bonds to maintain the triple helix also increases, thereby increasing the amount of
DMSO required to understand the triple helix.
After the triple helix is completely derotated, by reducing the mass fraction of DMSO/DMSO in water, the random coil can be reassembled into the triple helix, but the number of triple helix in the original state is reduced
.
However, DMSO, as a highly toxic organic solvent, poses a potential safety risk in the construction of complexes, so the construction process and products need to be rigorously monitored and tested
.

Self-assembly of CUR in alkali neutralization and treatment

It can be seen from Figure 5 that the NaOH concentration range of the triple helix polysaccharide with lower molecular mass maintains the triple helix conformation, indicating that compared with the polysaccharide with higher molecular weight, the lower molecular weight polysaccharide has more triple helix aggregates (THAs)
in NaOH solution.
Therefore, under alkali neutralization (AN) treatment, the influence of
THAs on the self-assembly of CUR should be fully considered.
In Fig.
5, all triple helix polysaccharides can maintain a relatively complete triple helix conformation at lower NaOH concentrations, and it is speculated that at this stage, the weak intermolecular hydrogen bonds in THAs are mainly destroyed, resulting in a large number of dissociation of THAs.
After the NaOH concentration is further increased, the strong intermolecular hydrogen bond between the three helical chains is destroyed, resulting in the unrotation of the monomeric triple helix (ITHs), and finally the formation of random coiling
.

The characteristics and limitations of self-assembly under the three induction methods are shown in
Table 1.

3.
Functional materials based on self-assembly of gel polysaccharides and their applications

Polymer self-assembly technology has made important contributions to the elucidating of the behavior of biological macromolecules and the construction of functional factor delivery systems, which is an important research project
in the field of food and materials science in recent years.
The nature of the reversible conformational transition of the triple helix (i.
e.
, the unwinding/re-spinning process) makes the triple helix polysaccharide exhibit unique molecular recognition behavior and incomparable characteristics, which can form an ordered hierarchy through self-assembly, and wrap various guest functional factors
with "supramolecular chemical packaging".
However, because polysaccharides do not have template synthesis characteristics such as DNA and proteins, polysaccharides are susceptible to differences
in structure, conformation, and biological activity due to molecular weight.
In view of this problem, the self-assembly of triple helix polysaccharides can spontaneously change the disordered sugar chain conformation into an ordered sugar chain conformation without changing the sugar chain composition, which can effectively reduce the conformational differences caused by the different molecular weights of polysaccharides, and at the same time improve the biological activity of polysaccharides and extend the functionality
of polysaccharides.

Conclusion