Metabolic microenvironment-forming porous hydrogels for the protection of phenylalanine ammonia-lyase in the intestine.

Biological therapeutics drive biomedical innovation, yet their oral delivery is hindered by gastrointestinal instability. Phenylalanine ammonia-lyase (PAL), a promising enzyme for phenylketonuria management, exemplifies this challenge. Here, we present ultraporous template-based hydrogels (TGels) that protect and deliver PAL in the small intestine. PAL-TGels are obtained via UV crosslinking of poly(ethylene glycol) diacrylamide in the presence of sacrificial CaCO3 templates, followed by absorption-based loading of PAL and a protease inhibitor. The interconnected porous matrix retains PAL while allowing phenylalanine and trans-cinnamic acid diffusion for continuous catalysis. In vitro studies confirmed robust protein retention and preserved enzymatic activity in simulated intestinal fluid, without impairing global trypsin function, while large animal tests validated matrix properties and transit. This fabrication-efficient platform enables localized intestinal enzyme protection, offering a promising route toward safe and accessible oral PAL therapy for phenylketonuria.