Dietary fiber (DF) was extracted from navel orange peel residue by enzyme (E-DF) and ultrasound-assisted deep eutectic solvent (US-DES-DF), and its physicochemical and prebiotic properties were characterized. Based on Fourier-transform infrared spectroscopy, all DF samples exhibited typical polysaccharide absorption spectra, indicating that DES could separate lignin while leaving the chemical structure of DF unchanged, yielding significantly higher extraction yields (76.69 ± 1.68%) compared to enzymatic methods (67.27 ± 0.13%). Moreover, ultrasound-assisted DES extraction improved the properties of navel orange DFs by significantly increasing the contents of soluble dietary fiber and total dietary fiber (3.29 ± 1.33% and 10.13 ± 0.78%, respectively), as well as a notable improvement in the values of water-holding capacity, oil-holding capacity, and water swelling capacity. US-DES-DF outperformed commercial citrus fiber in stimulating the proliferation of probioticBifidobacteriastrains in vitro. Overall, ultrasound-assisted DES extraction exhibited potential as an industrial extraction method, and US-DES-DF could serve as a valuable functional food ingredient. These results provide a new perspective on the prebiotic properties of dietary fibers and the preparation process of prebiotics.