Optimization of Proanthocyanidin Extraction from Grape Seeds Using Response Surface Methodology and Subsequent...

This study employed UPLC-MS/MS to determine the contents of major polyphenolic compounds and proanthocyanidins (PCs) in Kyoho grape seeds, optimized the extraction method and conditions for PCs using response surface methodology (RSM), and further evaluated the scavenging activities of PCs against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and hydroxyl (•OH) radicals as well as their effects on growth, immunity, and oxidative stress in mice. Three hundred and sixty 3-week-old male mice (42.28 ± 0.31 g) were assigned to a single factor complete randomized trial design and fed with six different diets including 0 mg/kg vitamin E(VE) + 0 mg/kg PCs, 100 mg/kg VE, 25 mg/kg PCs + 75 mg/kg VE, 50 mg/kg PCs + 50 mg/kg VE, 75 mg/kg PCs + 25 mg/kg VE and 100 mg/kg PCs, respectively. The results demonstrated that PCs were identified as the predominant phenolic compounds, accounting for 29.6% of total phenolic substances in Kyoho grape seeds. Additionally, the ultrasound-assisted extraction method was superior to the shaker-assisted and low-temperature infiltration extraction methods, with optimal conditions of 60% ethanol concentration, material-to-liquid ratio of 1:20 g/mL, temperature of 30 °C, and extraction time of 50 min. Scanning electron microscopy (SEM) revealed that ultrasound treatment effectively disrupted the seed surface structure, facilitating PC release. In vitro, PCs exhibited significantly stronger DPPH and hydroxyl radical (•OH) scavenging activities than vitamin C (VC), Trolox, and gallic acid. Compared with the control group, mice fed diets containing PCs and VE showed higher superoxide dismutase (SOD) activity, glutathione peroxidase (GSH-PX) activity, and total antioxidant capacity (TAOC), Catalase (CAT), GPX and inflammation factor 10 (IL-10) genes levels in the serum and liver (p < 0.05), whereas the levels of immunoglobulin G (IgG), immunoglobulin A (IgA), immunoglobulin M (IgM), tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6), as well as the mRNA expression of IL-1β and TNF-α, showed the opposite trend (p < 0.05). In conclusion, the antioxidant capacity of PCs was stronger than that of VC and VE. The addition of PCs improved the antioxidant activity and immune function of mice.