Evaluation of Molecular Assembly Morphology in Agarose Gel Using Depolarized Dynamic Light Scattering Method

The incorporation of molecular assemblies into hydrogels for transdermal delivery applications may affect their morphology and the dynamics of their constituent molecules; however, such effects have not been thoroughly investigated. In this study, we used bicellar mixtures composed of DPPC and DHPC as a model system to evaluate structural changes and reversibility within agarose hydrogels using depolarized dynamic light scattering (DDLS), a non-destructive method capable of analyzing morphologies of samples dispersed in an aqueous solution. To verify the performance of the DLS apparatus, standard particles and synthetic mica were measured, confirming that sufficient scattering intensity could be obtained even at an agarose concentration of 0.5 mass%. We investigated DPPC vesicles and DPPC/DHPC bicellar mixtures in aqueous solution and agarose gel, analyzing their translational and rotational diffusion coefficients. By applying a morphology-sensitive correction factor α, the aspect ratio (AR) of the bicellar assemblies was estimated to be approximately 2.0. The major and minor axes of the DPPC/DHPC bicelle mixtures were 40.0 nm and 20.0 nm in aqueous solution, and 32.7 nm and 15.0 nm in agarose gel, respectively. These differences may indicate stacking of multiple bicellar mixtures, which is consistent with the TEM image observations. In addition, although the structural transformation of bicellar mixtures in agarose gels occurred more slowly than in aqueous solution, changes induced by external stimuli (e.g., heating and stirring) and their partial reversibility were observed. These findings demonstrate that DDLS is a useful technique for evaluating the structural stability and responsiveness of molecular assemblies within agarose hydrogels, providing valuable insights for the future design of gel-embedded drug delivery systems.