Oral chitosan/alginate nanoparticles for enhanced bioavailability, intestinal retention, and sustained analgesic efficacy of NANT

Abstract Snake venom proteins have attracted considerable attention as oral analgesics owing to their definite analgesic effects and lack of tolerance or addiction. Herein, we report the successful development of an optimized oral analgesic nanoparticle formulation (CS/ALG@NANT NPs), prepared by the self-assembly of chitosan/alginate and Naja atra neurotoxin (NANT) via polyelectrolyte complexation and optimized using a systematic Box-Behnken design. The resulting nanoparticles exhibited uniform spherical morphology, an appropriate particle size (278 nm), high encapsulation efficiency (>90%), and substantial drug loading (>23%). CS/ALG@NANT NPs showed pH-responsive stability, maintaining structural integrity under gastric conditions while enabling rapid intestinal release, as well as excellent cytocompatibility and time-dependent cellular uptake. In vivo imaging revealed strong intestinal targeting, prolonged retention, and minimal off-target distribution, thereby protecting NANT from gastric degradation. Pharmacodynamic studies demonstrated significant dose-dependent analgesic effects, with high-dose efficacy comparable to reference drugs. These findings indicate that CS/ALG@NANT NPs represent a promising oral delivery platform for sustained analgesia and enhanced intestinal targeting, offering strong potential for clinical pain management.