Immunomodulatory activity of egg yolk protein hydrolysates prepared by novel two-step hydrolysis: A study of mechanism and stability after in vitro digestion model.

The aim of this study was to determine the immunomodulatory activity of 2-step egg yolk protein hydrolysates. A two-step hydrolysate of egg yolk protein was prepared using 2 enzymes sequentially, pancreatin and neutrase (EYPH-PN). Our results illustrated that EYPH-PN increased the expression of inducible nitric oxide synthase (iNOS) mRNA in macrophages, resulting in increased nitric oxide (NO) production. EYPH-PN could also enhance the production of tumor necrosis factor (TNF)-α and interleukin (IL)-6 at both the mRNA and protein levels in macrophages. In addition, treatment with EYPH-PN increased the phagocytic activity of macrophages. According to the evaluation with specific inhibitors, both p38 and JNK cell signaling pathways were involved in the activation of macrophages induced by EYPH-PN. As the TLR-2 receptor of macrophages was blocked, the NO production induced by EYPH-PN was decreased. These results suggest that EYPH-PN activates RAW 264.7 macrophages via the TLR-2/p38/JNK pathway to increase the production of NO, TNF-α, and IL-6, and increases phagocytic activity. Furthermore, the immunomodulatory activity of EYPH-PN was maintained even after applying the in vitro digestion model. Taken together, EYPH-PN could be used as a functional food ingredient with excellent immunomodulatory activity in the food industry. Therefore, this study suggests a new alternative method to effectively utilize egg yolk protein, a by-product of the poultry industry.