Effect of Wall-Material Assembly Sequence on Ovalbumin–Chitosan Nanoparticles for Antarctic Krill Peptide Delivery

The objective of this study was to explore the effect of the assembly sequences of wall materials on the structure and properties of Antarctic krill peptide (AKP)-loaded ovalbumin (OVA)-chitosan (CS) nanoparticles (NPs). Two AKP-loaded NPs (CS/OVA-AKP and OVA/CS-AKP) were prepared by changing the sequences of OVA and CS. The results confirmed that CS/OVA-AKP had a smaller particle size (291 nm vs. 320 nm), lower polydispersity index (0.233 vs. 0.282), higher absolute zeta potential (34.4 mV vs. 32.1 mV), and higher encapsulation efficiency (81.6% vs. 75.4%) than OVA/CS-AKP. X-ray diffraction analysis confirmed that AKP was encapsulated in an amorphous state within the NPs. Fourier transform infrared spectroscopy and three-dimensional (3D) fluorescence spectroscopy revealed that electrostatic interactions, hydrogen bonding, and hydrophobic interactions were the primary driving forces for nanoparticle formation, with CS/OVA-AKP demonstrating a stronger OVA fluorescence quenching effect. Compared with OVA/CS-AKP, CS/OVA-AKP exhibited better redispersibility, and CS/OVA-AKP showed greater stability under various environmental factors (thermal treatment, salt concentration, pH, and storage time). During simulated gastrointestinal digestion, CS/OVA-AKP effectively protected AKP from gastric degradation and showed a higher AKP release rate in simulated intestinal fluid (61.1%) than OVA/CS-AKP (53.0%). The release followed the Korsmeyer-Peppas model, with OVA/CS-AKP exhibiting non-Fickian diffusion (<i>n</i> = 0.7500), and CS/OVA-AKP approached Case II transport (<i>n</i> = 0.9889), indicating erosion-controlled release behavior. CS/OVA-AKP also demonstrated higher hypoglycemic activity, with inhibition rates of 41.1%, 37.5%, and 36.1% for α-glucosidase, α-amylase, and DPP-IV, respectively. These findings underscore the important influence of wall-material assembly sequences on the structure and properties of AKP-loaded NPs, offering valuable insights for the development of bioactive peptide delivery systems.