(Invited) Elucidating the Micro Phase-Separation of Mixed Kappa and Iota Carrageenan Gels Using Multiple Particle Tracking
Authors: Geonzon Lester Canque , Bacabac Rommel Gaud , Matsukawa Shingo 
Published: February 2020

Carrageenan is a family of linear sulfated polysaccharides extracted from seaweeds. Among the different types of carrageenan, kappa carrageenan (KC) and iota carrageenan (IC) are widely used in industries due to its gelling behavior. Because of its importance in industries, KC and IC as well as its mixtures have been extensively studied using different methods. However, it is still unclear whether mixed KC:IC undergo a phase separated network structure of IC-rich and KC-rich or interpenetrated network structure. In this report, the network structure and gelation mechanism of KC, IC and its mixture were elucidated using multiple particle tracking.
Sodium-type κ- and ι- carrageenan powder was purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan). All samples were dialyzed against NaCl solution and subsequently against deionized water, to obtain Na+ type carrageenan solutions. Samples were prepared by diluting carrageenan and KCl solutions and heated for 30 min at 70 °C under vigorous stirring. The prepared solutions were mixed using vortex to obtain 1.5% w/w carrageenan with 10 mM KCl and heated at 90 °C for 20 min. Fluorescently labelled 0.1 μm particles (Green, ThermoScientific) were added at a concentration of ~0.01% (v/w) at 90 °C and stirred for another 5 min to assure homogenous dispersion. Mixture of KC and IC was prepared at different volume fraction of IC. Movies of the diffusing particles were recorded at a frame rate of 7.5 frames per second for 110 seconds. In cooling experiments, particle tracking was performed at different sample temperatures from 50 °C to 10 °C. In storage experiments, hot solutions of carrageenan were cooled down from 50 °C to 5 °C at a rate of 1 °C/min in a temperature-controlled incubator to form a gel. The gelled sample was stored at 5 °C prior to experiment and particle tracking was performed at 5 °C. Particle tacking was implemented in Mathematica 10 (Wolfram Research, Inc., USA).
The temperature dependence of the mean square displacement (MSD) of particles in KC, IC and mixed KC:IC gels were carried out to quantify the dynamics of the evolving network structure on cooling as shown in Figure 1. In mixed KC:IC, a two-step decrease in MSD was observed. The onset of the observed decrease is close to the decrease in pure KC and IC gels. This trend is consistent with the observed increase in the viscoelastic modulus in dynamic viscoelasticity measurements. The decrease in the MSD with decreasing temperature is due to the steric hindrance created by the formation of clusters of IC and KC chain aggregates. This suggests that KC and IC chains formed aggregates independently on cooling. In pure carrageenan, IC gels formed weak entanglement among clusters of IC chain that allowed diffusion of particles through crevices of the entangled clusters while KC gels rapidly formed thick aggregates of interconnected clusters that confined the particles. On the other hand, in mixed KC:IC, the presence of IC clusters decreased the mobility of KC chains at low temperature thus slowed the formation of stable clusters of KC.
With storage time, a broad distribution of MSD was observed for the mixed gels. This suggests that an inhomogeneous solution was formed as KC and IC clusters continue to rearrange to form a stable network. The degree of heterogeneity in the mixed gels is reflected in the corresponding van Hove distribution of particle displacement and the distribution of MSD as a function of slope. In the van Hove distribution, a bimodal Gaussian form with a broad and narrow Gaussian width fit well on the distribution suggests a mobile and immobile fraction of particle at each mixing ratios. In the distribution of MSD as a function of slope, the mobile and immobile particles were identified and clustered accordingly. This suggests that the presence of mobile particles was due to the existence of IC-rich domains that allowed diffusion of particles and the immobile particles was due to the confinement of particles in KC-rich domains as shown in the graphical representation in Figure 1. This is consistent with the proposed gelation mechanism of a local phase separation into IC-rich and KC-rich domains. As the volume fraction of IC was increased, the number of mobile particles increased consequently and suggests a phase transition from a sea-island (80%KC:20%IC) to a bi-continuous (50%KC:50%IC) to an island-sea (30%KC:70%IC) network structure is likely to occur.

Figure 1