Sequential extraction, structural characterization, and biological activities of polysaccharides from olive (Olea europaea L.) pomace.

Olive pomace, a by-product of olive oil production, remains underexplored despite its potential environmental and economic benefits. This study sequentially extracted three polysaccharides (OERC, OERH, and OERA) from olive pomace using water at room temperature, hot water (80 °C), and 2% Na₂CO₃ solution (60 °C). Characterization through UV-Vis spectroscopy, Fourier transform infrared spectroscopy, scanning electron microscopy, thermogravimetric analysis, high-performance liquid chromatography, and Congo red experiment revealed that these polysaccharides are heteropolysaccharides primarily composed of glucose, mannose, rhamnose, and galactose, each with distinct molar ratios. The polysaccharides exhibited significant antioxidant activity by scavenging hydroxyl, DPPH, and superoxide radicals, with OERC showing the highest potency. They also repaired H₂O₂-induced oxidative damage in HepG2 cells, with OERC at 400 μg/mL and OERH/OERA at 200 μg/mL displaying optimal efficacy. Additionally, these polysaccharides effectively inhibited α-glucosidase activity, enhanced glucose consumption, and increased glycogen content in insulin-resistant models, thereby exerting hypoglycemic effects. Furthermore, they promoted proliferation, phagocytosis, and nitric oxide release in RAW264.7 macrophages, demonstrating immunomodulatory effects. These results indicate that sequential extraction under varying conditions is an effective method for preparing polysaccharides. The three polysaccharides isolated in this study show great potential for development as functional products with antioxidant, hypoglycemic, and immunomodulatory applications.