This in vitro study compared the cutting efficiency of natural diamond burs (NDB) and synthetic diamond burs (SDB) on zirconia and natural teeth. Two types of diamond rotary instruments with identical specifications (TR-13, Tapered Round End) were evaluated using 100 zirconia specimens and 100 extracted molars. Each bur was tested through 10 consecutive cutting cycles under standardized conditions (200,000 rpm, 2 N force, water coolant). Cutting efficiency was evaluated based on weight loss per unit time (mg/min). Surface characterization was performed using field-emission scanning electron microscopy (FE-SEM) and energy-dispersive X-ray spectroscopy (EDS). For zirconia cutting, NDB demonstrated significantly higher cutting efficiency than SDB at all cycles (p < 0.05), with 38.9% greater total material removal (115.97 ± 2.22 mg versus 83.46 ± 2.08 mg). NDB also exhibited lower wear rates (40.00% versus 49.68% reduction). For natural tooth cutting, no significant differences were observed between NDB and SDB (p > 0.05). EDS analysis showed that SDB exhibited greater carbon loss (13.46% versus 2.49%) and increased surface heterogeneity after repeated use. These findings indicate that natural diamond burs are superior for cutting high-hardness materials such as zirconia, while both bur types perform equivalently on natural teeth.