Abstract
Oyster peptides (OPs) have gained increasing attention for their excellent biological activities, especially immunomodulatory effects. In this study, oyster proteins were fermented using Lactobacillus casei to prepare bioactive peptides, and the effects of fermentation parameters (time, temperature, and inoculum amount) on the degree of hydrolysis (DH) were optimized. The optimal fermentation conditions were determined as 30 h, 35 °C, and 5% inoculum amount, resulting in a DH of 28.24%. Structural characterization showed that OPs were mainly composed of low-molecular-weight peptides (<1000 Da) with high hydrophobic amino acid content, and they exhibited good stability during in vitro gastrointestinal digestion. In vitro immunological evaluation using RAW264.7 macrophages demonstrated that OPs significantly enhanced phagocytic activity and nitric oxide (NO) production, and upregulated the mRNA expression levels of pro-inflammatory cytokines including interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α. Mechanistically, OPs exerted immunostimulatory effects by specifically activating the extracellular signal-regulated kinase (ERK) pathway within the mitogen-activated protein kinase (MAPK) signaling cascade, without significant alterations in the phosphorylation levels of p38 and c-Jun N-terminal kinase (JNK). These findings highlight the potential of Lactobacillus casei-fermented oyster peptides as natural immunomodulatory ingredients for functional food development.