To investigate the root morphology and root canal anatomy of maxillary first premolar using microscopic computed tomography (micro-CT).324 maxillary first premolars were collected and scanned. The root and canal diameter, canal wall thickness, root taper, and cross-sectional shapes were determined in the single root with 1 canal (SR1C), single root with 2 canals (SR2C), and 2 roots with 2 canals (2R2C) by micro-CT.The results showed that single-rooted maxillary premolars were more common than other types. The incidence of SR1C, SR2C, and 2R2C reached 25%, 26.39%, and 26.39%, respectively. Root and canal diameters and canal wall thickness were decreased from coronal third to apical foramen. The three parameters and canal taper showed increases from buccal and palatal (BP) to mesiodistal (MD) aspects. The root canal tapers were smallest of the middle third level. The findings showed the different variations in 2R2C teeth. The root canal cross-sectional morphology in maxillary first premolars is complicated, including round, oval, long oval, flat canal, and irregular canal shapes. The distribution varied in different aspects.Root canal morphology showed a wide variation and complicated structure. The single-rooted teeth were more common in the Chinese adolescent population, and the majority of maxillary first premolars have two canals.