Ionic modulation of IPN formation and recovery in hyaluronic acid/κ-carrageenan hydrogels: Insights from echo-DWS microrheology, bulk rheology and FTIR 

Local confinement arising from polymer chain mobility in polysaccharide-based hydrogels can be directly correlated with the formation of interpenetrating polymer networks (IPNs) which can be modulated via ion charge density, and subsequently governs their mechanical properties for injectable applications. Here, we establish a multi-scale framework that integrates echo-diffusing wave spectroscopy (echo-DWS) microrheology with FTIR spectroscopy and bulk rheology to provide a mechanistic bridge between molecular interactions, resulting network architecture and bulk mechanical recovery. Specifically, we show that microrheology captures the transition from diffusive to sub-diffusive motion induced by the presence of ions; we also see a distinct microrheological signature with multivalent ions which we propose is a predictor of formation of semi-interpenetrated networks in a hyaluronic acid/kappa-carrageenan hydrogel system. Probing further with FTIR spectroscopy, we incorporate ionic strength as an additional lever to validate our findings on ion charge density from microrheology- K+ did not produce detectable sulfate or carboxylate shifts, consistent with a predominantly physical mixture and the absence of IPN formation while Ca2+ induced concentration-dependent shifts in both the amide/carboxylate region and the sulfate band, consistent with local chain compaction, complexation of HA carboxylates and bridging between κ-CG helices, indicating promotion of a semi-interpenetrated network structure above a threshold concentration. Al3+ induced a distinct shoulder in the HA carboxylate region, confirming HA coordination and co-crosslinking with κ-CG, possibly yielding a semi-IPN. Finally, we investigate mechanical recovery as a function of both ion charge density and strength, to understand how these critical design levers may be leveraged in tailoring mechanical properties of supramolecular, interpenetrating network hydrogels for injectable applications.