The purpose of this study is to assess segmentation reproducibility of artificial intelligence-based algorithm, TotalSegmentator, across 34 anatomical structures using multiphasic abdominal CT scans comparing unenhanced, arterial, and portal venous phases in the same patients. A total of 1252 multiphasic abdominal CT scans acquired at our institution between January 1, 2012, and December 31, 2022, were retrospectively included. TotalSegmentator was used to derive volumetric measurements of 34 abdominal organs and structures from the total of 3756 CT series. Reproducibility was evaluated across three contrast phases per CT and compared to two human readers and an independent nnU-Net trained on the BTCV dataset. Relative deviation in segmented volumes and absolute volume deviations (AVD) were reported. Volume deviation within 5% was considered reproducible. Thus, non-inferiority testing was conducted using a 5% margin. Twenty-nine out of 34 structures had volume deviations within 5% and were considered reproducible. Volume deviations for the adrenal glands, gallbladder, spleen, and duodenum were above 5%. Highest reproducibility was observed for bones (− 0.58% [95% CI: − 0.58, − 0.57]) and muscles (− 0.33% [− 0.35, − 0.32]). Among abdominal organs, volume deviation was 1.67% (1.60, 1.74). TotalSegmentator outperformed the reproducibility of the nnU-Net trained on the BTCV dataset with an AVD of 6.50% (6.41, 6.59) vs. 10.03% (9.86, 10.20; p < 0.0001), most notably in cases with pathologic findings. Similarly, TotalSegmentator’s AVD between different contrast phases was superior compared to the interreader AVD for the same contrast phase (p = 0.036). TotalSegmentator demonstrated high intra-individual reproducibility for most abdominal structures in multiphasic abdominal CT scans. Although reproducibility was lower in pathologic cases, it outperforms both human readers and a nnU-Net trained on the BTCV dataset.