Abstract: To investigate the structure of Alpinia oxyphylla Miq. oligosaccharides (MC) and their impact on human gut microbiota, MC was extracted and purified using ultrasound-assisted high-voltage pulsed electric fields. Its structural characteristics were analyzed using Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and scanning electron microscopy (SEM). Its in vitro antioxidant and hypoglycemic activities, as well as its regulatory mechanism on the intestinal flora were investigated. Results showed that the extracted MC was a glucose homopolymer (100% glucose monosaccharide composition) with a molecular weight of 2461 Da. The β-glycosidic bond configuration of MC was confirmed by FT-IR spectroscopy, with characteristic peaks observed at 1656 cm⁻¹ and 1623 cm⁻¹. A hierarchical structure characterized by coexisting crystalline and amorphous domains within the β-glucan backbone was demonstrated through XRD and SEM analyses. These structural features constitute a structural basis for targeting gut microbiota. In vitro activity assays indicated that MC exhibited negligible antioxidant activity but demonstrated notable hypoglycemic activity. Fecal fermentation experiments demonstrated that MC markedly increased the abundance of Firmicutes, reduced Bacteroidetes, and enriched the genera Ligilactobacillus and Weissella. These effects are potentially mediated by β-glycosidic bond stability prolonging intestinal retention time, and amorphous region porosity enhancing bacterial adhesion efficiency to remodel microbial balance. This study provides a theoretical foundation for developing Alpinia oxyphylla Miq. oligosaccharides as prebiotics.