Ion valency-driven control of network architecture and tissue-level drug transport in protein-derived hydrogels

Controlling molecular transport in hydrated macromolecular networks remains a central challenge in the development of protein-based drug delivery systems. Herein, we report a biocompatible human serum albumin (HSA)-derived hydrogel that spontaneously self-assembles at room temperature through simple mixing with a reducing agent, tris(2-carboxyethyl)phosphine (TCEP), and inorganic salts, without the use of synthetic crosslinkers. Reduction of intramolecular disulfide bonds induces partial unfolding of HSA, promoting intermolecular association and network formation. By systematically varying the valency of inorganic cations, we observed consistent modulation of hydrogel gelation behavior, mechanical properties, and apparent network characteristics. The results are consistent with a proposed mechanism in which monovalent cations mainly reduce electrostatic repulsion between protein chains, whereas divalent cations may promote stronger ion-mediated associations, leading to relatively denser and mechanically reinforced networks. Using doxorubicin as a model small-molecule probe, we show that molecular release behavior is associated with cation-dependent differences in hydrogel properties, resulting in tunable release profiles of approximately 37-50% within 24 h without relying on specific drug-matrix affinity. In vitro release and ex vivo permeation studies further suggest that the representative HSA hydrogel can exhibit tissue-dependent permeation and retention behavior, supporting its potential as a localized drug delivery platform. These findings establish a clear structure-property-transport relationship in protein-derived hydrogels and provide a simple ionic strategy for designing tunable and cytocompatible hydrogel platforms for localized and transdermal drug delivery.