The accuracy of tooth segmentation in intraoral scans is crucial for performing virtual setups and appliance fabrication. Hence, the objective of this study was to estimate and compare the accuracy of automated tooth segmentation generated by the artificial intelligence of dentOne software (DIORCO Co, Ltd, Yongin, South Korea) and Medit Ortho Simulation software (Medit Corp, Seoul, South Korea). Twelve maxillary and mandibular pretreatment dental scan sets comprising 286 teeth were collected for this investigation from the archives of the Department of Orthodontics, Faculty of Dentistry, Alexandria University. The scans were imported as standard tessellation language files into both dentOne and Medit Ortho Simulation software. Automatic segmentation was run on each software. The number of successfully segmented teeth vs failed segmentations was recorded to determine the success rate of automated segmentation of each program. Evaluation of success and/or failure was based on the software's identification of the teeth and the quality of the segmentation. The mesiodistal tooth width measurements after segmentation using both tested software programs were compared with those measured on the unsegmented scan using Meshmixer software (Autodesk, San Rafael, Calif). The unsegmented scans served as the reference standard. A total of 288 teeth were examined. Successful identification rates were 99% and 98.3% for Medit and dentOne, respectively. Success rates of segmenting the lingual surfaces of incisors were significantly higher in Medit than in dentOne (93.7% vs 66.7%, respectively; P<0.001). DentOne overestimated the mesiodistal width of canines (0.11 mm, P= 0.032), premolars (0.22 mm, P< 0.001), and molars (0.14 mm, P= 0.043) compared with the reference standard, whereas Medit overestimated the mesiodistal width of premolars only (0.13 mm, P= 0.006). Bland-Altman plots showed that mesiodistal tooth width agreement limits exceeded 0.2 mm between each software and the reference standard. Both artificial intelligence-segmentation software demonstrated acceptable accuracy in tooth segmentation. There is a need for improvement in segmenting incisor lingual tooth surfaces in dentOne. Both software programs tended to overestimate the mesiodistal widths of segmented teeth, particularly the premolars. Artificial intelligence-segmentation needs to be manually adjusted by the operator to ensure accuracy. However, this still does not solve the problem of proximal surface reconstruction by the software.