P26S

23AG011

BL-12

Monitoring the Self-Assembly of Alginate Hydrogel Using
Synchrotron Radiation Circular Dichroism
Tatsuki Hagaa, Masaya Yoshida, Takeharu Hainoa, Koichi Matsuob andMohamed I.A. IbrahimbaGraduate School of Advanced Science and Engineering, Hiroshima University.
Hiroshima Synchrotron Radiation Center (HiSOR), Hiroshima University, 2-313 Kagamiyama, HigashiHiroshima, Hiroshima 739-0046 Japan.

b

Keywords: Alginate, Hydrogel, Self-Assembly, CD, AFM

Circular dichroism (CD) spectroscopy, traditionally used for studying the secondary structure of proteinsand peptides [1], has not been yet established for studying polysaccharides (PSs). The current study has
explored the potential of using CD in structural changes associated with PS hydrogelation. Certain PSs canchange from a liquid to a gel when exposed to light, ions, or temperature. Alginate, an ecofriendly andnontoxic PS used in medicine and food, is especially known for forming hydrogels when mixed with
polyvalent ions [2]. In the food industry, alginate serves as an additive to enhance texture and shape. Thisfunction is closely related to the conformational changes during sol-gel transitions, so structural analysis ofalginate is helpful for maximizing its function as an additive.
Herein, CD experiments using synchrotron radiation (SR) light were carried out to investigate thestructural changes associated with gel formation in the presence and absence of metal ions. Monovalent ionssuch as Na+, K+, and Li+ did not induce hydrogel formation, whereas hydrogels were obtained by divalents(e.g., Ca2+, Ba2+, Cu2+, and Zn2+), as well as trivalent ion (e.g., Fe3+). The cations exhibited different
influences on the native conformation of alginate, suggesting variations in the assembly of the alginate chainsin the presence of different ions. Additionally, discrepancies in CD spectra reflected that the formedhydrogels possessed diverse mechanical, physical properties, and morphology.
The formulation of hydrogels is an evidence of the self-assembly phenomeneon. Alginate chains are interconnected, forming a cross-linked 3D-network structure that entraps water molecules, leading to hydrogel
formation. Hence, to construct the self-assembly/gel phase diagram based on the CD observations,concentration-dependent experiments, varying alginate concentrations or polyvalent ion concnetrations (i.e.,Ca2+), were conducted using a CD spectrometer. The concentration of the polyvalent ion that causes nochange in the CD spectrum of alginate or that self-assembly of alginate chains was terminated (Figure 1,left) was marked as the initiation of aggregation. As a result, a Ca2+ concentration of 1.56 mM, at 1.0 mg/ml
alginate concentration, was referred to the transition concentration from the self-assembly to aggregation
which is followed by hydrogel formation (Figure 1, right). This finding was reinforced by atomic force
microscopy (AFM) imaging, which showed increased cross-linking and thichness of fibers at higher Ca2+
concentration (2.5 mM) (Figure 2).

FIGURE 1. CD spectrum of alginate (1.0 mg/ml) as a function of Ca2+ concentrations (0.5 mM ~ 2.5 mM) (left), and selfassembly transition diagram based on CD observations (right).

FIGURE 2. AFM images of the alginate-Ca2+ system with [alginate] = 1.0 mg/ml and Ca2+ concentrations of 1.0 mM (left),or 2.5 mM (right).

Ongoing investigations are underway to further explore the alginate-Ca2+ hydrogel system using SRCD,
aiming to study the kinetics and thermodynamics of hydrogelation process.

REFERENCES[1] K. Matsuo, G. Kunihiko, Bulletin of the Chemical Society of Japan, 2013, 86, 675-689.
[2] J. Brus, M. Urbanova, J. Czernek, Biomacromolecules, 2017, 18, 2478–2488.