Stabilization mechanisms, cell density, and survival of probiotic encapsulation prepared with thiolated hyaluronic acid-whey protein complex: For potential colon targeting probiotic delivery system.

To improve the stress resistance of Lactococcus lactis subsp. lactis (L. lactis) KLDS 4.0325 in harsh environments, we constructed a novel thiolated hyaluronic acid (HASH)-whey protein isolate (WPI) encapsulation system. The formation of a non-covalent HASH-WPI complex was confirmed via Fourier-transform infrared spectroscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incrementing the amount of HASH in the complex significantly increased the particle size and absolute Zeta potential to 1393.67 ± 44.15 nm and 53.28 ± 0.68 mV, respectively, while instilling excellent antioxidant activity: 28.41 ± 0.51% scavenging of 2,2-diphenyl-1-picrylhydrazyl and 41.80 ± 0.86% scavenging of 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid). In addition, increasing the HASH content also caused a transition of the secondary structure of WPI from α-helix to β-sheet, with the latter content increasing from 20% to 24%. Morphological analysis shows that the 0.8% HASH-WPI complex exhibited a uniformly dispersed and compact structure that provided the best protective effect for L. lactis KLDS 4.0325, with survival rates in 0.8% HASH-WPI microcapsules of 96.04 ± 0.49% after freeze-drying, 76.58 ± 0.13% after gastrointestinal digestion, and 93.12 ± 0.24% after pasteurization. This study confirms that the dense network structure formed by the HASH-WPI complex can serve as a physical barrier to provide enhanced protection for probiotics. Additionally, ex vivo intestinal imaging directly demonstrated that the complex significantly increased probiotic adhesion to the colon. This study provides a promising strategy for constructing efficient, stable, and colon-retentive probiotic delivery systems.