Digested casein-stabilized co-dissolution of calcium and iron in oxalate system for enhanced metal bioavailability

Oxalate severely impairs calcium and iron bioavailability by forming insoluble precipitates in the gastrointestinal tract, posing challenges for mineral nutrition in plant-based diets. This study investigated the thermodynamic and kinetic interactions between calcium, iron, and oxalate, and explored whether digested casein could enhance mineral bioavailability under oxalate challenge. Thermodynamic analysis revealed that iron(II) oxalate exhibited higher solubility product (K_sp) and weaker association (K_a = 4655) compared to calcium oxalate (K_a = 25126) at 25 ℃. Conductometric titration demonstrated that iron salts delayed calcium oxalate precipitation through competitive oxalate binding and cluster stabilization, with iron(II) gluconate showing better effects. Kinetic studies identified calcium lactate paired with iron gluconate as the optimal combination, maintaining iron solubility over 2 h in oxalate-rich conditions. Digested casein, prepared under simulated gastrointestinal conditions, dose-dependently enhanced calcium and iron solubility by forming stable peptide-mineral-oxalate nanoparticles, characterized by transmission electron microscopy and dynamic light scattering. These nanoparticles exhibited either crystalline (inorganic salts) or amorphous (organic salts) structures depending on counter-ion identity. Critically, Caco-2 cell studies demonstrated that digested casein-mediated nanoparticles restored both calcium and iron absorption despite oxalate presence, operating primarily through caveolin-mediated endocytosis rather than classical mineral transporters. This work reveals a novel mechanism whereby digested casein neutralizes oxalate's antinutritional effects through nanoparticle formation, providing a promising nutritional strategy for enhancing mineral bioavailability in oxalate-rich diets.