Abstract Two experiments were conducted to investigate the kinetics of phosphorus (P) absorption and expressions of type IIb sodium‐dependent phosphate cotransporter (NaP‐IIb), inorganic phosphate transporters 1 and 2 (PiT‐1 and PiT‐2) in primary cultured duodenal epithelial cells of chick embryos. In experiment 1, the P absorptions across duodenal epithelial cell monolayers at different incubation time points (0, 10, 20, 40, 60, 80, 100 and 120 min) were compared. In experiment 2, the kinetics of P absorption was performed at 40 min after incubation of duodenal epithelial cells with the media containing 0, 0.75, 1.5, 3.0, 6.0, 12.0, 24.0 and 48.0 mmol P/L as KH 2 PO 4 , and the mRNA and protein expression levels of NaP‐IIb, PiT‐1 and PiT‐2 in duodenal epithelial cells with the media containing 0, 6.0 and 48.0 mmol P/L were determined at 87 min after incubation. The results from experiment 1 showed that the P absorption increased linearly ( p < .0001) from 0 to 80 min and the fastest increase occurred at 40 min; the asymptotic model was shown to have the best fit degree, and the optimal incubation time for saturable P absorption was determined to be 87 min. The kinetic curves of P absorption from experiment 2 demonstrated that P absorption was a mixed process of a non‐saturable diffusion plus a saturable carrier‐mediated transport across the duodenal epithelial cells. The high P concentration (48.0 mmol/L) decreased ( p < .05) NaP‐IIb and PiT‐1 mRNA and protein levels and increased ( p < .0001) PiT‐2 mRNA level. These results indicated that the P absorption across primary cultured duodenal epithelial cell monolayers of chick embryos was a mixed process of a non‐saturable diffusion plus a saturable carrier‐mediated transport and could be restricted by reducing the NaP‐IIb and PiT‐1 expressions while increasing the PiT‐2 expression at a high P concentration.