Nano-calcium phosphate (nCaP) has potential applications in nanomedicine fields such as drug delivery, bioimaging, antibacterial treatment, and bone formation promotion. However, its distribution and metabolic patterns within the body are not yet fully understood and require further in-depth research. This study employs a rare earth europium ion fluorescence labeling method and uses tumor-bearing mice as a model to investigate the distribution and metabolism of two sizes of nCaP (nanodots NDs: (2.53±0.63) nm; nanoparticles NPs: (107.76±25.37) nm×(17.66±1.63) nm) in the liver, spleen, lung, kidney, and tumor tissue. The results showed that after tail vein injection of 200 μL with a mass concentration of 1.5 mg/mL nCaP into tumor-bearing nude rats for 4 h, CaP NPs were primarily distributed in the liver and spleen, accounting for 65.70% and 29.32%, respectively, with 3.83% in the lung, while only 0.84% and 0.32% in the kidney and tumor. This suggests that larger CaP NPs are more easily captured by phagocytes within the reticuloendothelial system (RES). In contrast, compared to CaP NPs, accumulation of CaP NDs in the liver, spleen, and lung decreased significantly by 89.40%, 87.00%, and 88.89%, respectively, while their accumulation in the kidney and tumor increased by 3.67 and 3.06 times. This indicates that smaller particle size facilitates CaP NDs in glomerular filtration for urinary excretion and enhances their tumor-targeting capability. The clearance rates (CLz) of CaP NDs in the liver, spleen, and lung were 6.60, 4.14, and 2.40 times higher than that of CaP NPs, respectively, and 42.29% in the kidney. This indicates that reduced size of CaP NDs facilitates rapid metabolism by phagocytes in the liver, spleen, and lung but also results in reabsorption in the renal tubules. In tumor, the CLz of CaP NDs decreased by 91.9%, much smaller than that of CaP NPs, suggesting that the smaller CaP NDs exhibit significantly enhanced tumor targeting and retention capability. In the meantime, a physiologically based pharmacokinetic (PBPK) model incorporating particle size factors was preliminarily established for tumor-bearing mice to simulate the distribution of nano-calcium phosphate. The model's predictive fit (R²) for CaP NDs and CaP NPs in tumor sites reached 0.925 and 0.827, respectively. This study provides promising support for understanding in vivo distribution and metabolic patterns of nCaP and applying potential in medicine.