Antioxidant and Immune Enhancement Effects of <i>Zophobas atratus</i> Larvae Low-Temperature Extract on Macrophages through MAPKs and NF-κB Signaling Pathways

This study evaluated the protein content, antioxidant activity, and immune-enhancing potential of Zophobas atratus larvae extract (ZAE) and assessed its potential as a functional material for immune enhancement.The ZAE extraced at 5C (5C ZAE) had a higher protein content than the 70C extract, according to a bicinchoninic acid (BCA) assay.In addition, the 5C ZAE exhibited superior antioxidant activity in exhibited 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and ferric reducing antioxidant power (FRAP) assays.When ZAE was applied to RAW 264.7 cells, neither extract showed cytotoxicity, but the 5C ZAE resulted in a higher survival rate, Accordingly, subsequent experiments were carried out using the 5C ZAE.The secretion of cytokines (TNF-, IL-6) and nitric oxide (NO) increased in a concentration-dependent manner, as confirmed by ELISA and NO assays, and the expression levels of iNOS/COX-2 were also elevated.Furthermore, the 5C ZAE promoted immune activity through MAPKs phosphorylation and the activation of the NF-B pathway.These findings suggest that the low-temperature extract of Zophobas atratus larvae can be utilized as a functional material with both antioxidant and immune-enhancing effects.