Living drug carriers, including probiotics, prebiotics, and engineered microbes, are reshaping the field of therapeutic delivery by providing dynamic, host-responsive platforms with the capacity to address complex gastrointestinal, metabolic, neurological, and immune disorders. Encapsulation has emerged as a powerful approach to improve probiotics stability and performance by providing physical and biochemical barriers against stress factors. Conventional methods, such as spray-drying, extrusion, or emulsification, can improve survival, but often at the expense of reduced cell viability or limited scalability. Unlike bead- or film-forming processes, cryogelation generates interconnected macroporous networks under sub-zero conditions, allowing high cell loading while maintaining metabolic exchange and recovery of viable cells. These features are particularly advantageous for living drug carriers, as they combine effective protection with structural properties that support the long-term persistence and functional activity of encapsulated microbes.

This study investigated the structural, functional, and biological performance of spirulina (SPI), chlorella protein isolate (CPI), and blended SPI:CPI, sodium alginate (NaAlg) cryogels as delivery systems for Lacticaseibacillus rhamnosus GG (LGG). FTIR confirmed characteristic amide and carbohydrate-associated bands, while deconvolution of the amide I region indicated protein aggregation dominated by intermolecular β-sheet structures. Water sorption analysis revealed type III isotherms with high monolayer moisture contents, attributable to the hygroscopic maltodextrin–protein–alginate matrix. Thermogravimetric analysis showed four major mass-loss events corresponding to residual moisture, glycerol decomposition, polysaccharide breakdown, and protein–carbohydrate degradation. Mechanical testing indicated comparable stiffness and hardness across all formulations, suggesting that structural integrity is governed primarily by ice-templated macroporosity. μCT and SEM visualisation confirmed highly porous (76–78%) cryogel networks with protein-dependent differences in pore-wall smoothness, thickness, and collapse behaviour under high humidity. All cryogels disintegrated rapidly upon hydration, consistent with capillary-driven infiltration and erosion. Semi-dynamic gastrointestinal digestion demonstrated distinct colloidal transitions, with SPI forming compact gastric aggregates that improved LGG retention and controlled release in the intestine. Intestinal viability was highest for SPI cryogels, which also preserved adhesive capacity toward intestinal epithelium, indicating maintenance of key surface structures. Weibull modelling of storage data revealed strong effects of water activity and temperature on LGG survival. At aw 0.11 and 20 °C, shelf-lives ranged from 183–320 days, whereas high humidity or elevated temperature drastically accelerated inactivation. Across conditions, β values >1 indicated convex inactivation curves associated with cumulative cellular damage. Overall, SPI-based cryogels provided the most robust protection during processing, storage, digestion, and epithelial adhesion, highlighting their potential as effective protein-based carriers for controlled probiotic delivery.

Conference Theme: Innovative hydrocolloid design for delivering optimal nutrition and functional foods