To characterize the relationship between the cFGF23/Klotho ratio and phosphate level in patients with chronic kidney disease (CKD). A total of 152 patients with CKD stage 3-5 (CKD stage 3: n = 74; CKD stage 4: n = 60; CKD stage 5: n = 18) were included in the study. Thirty healthy volunteers served as controls. Intact-FGF23, cFGF23, Klotho, serum calcium, serum phosphate, and serum creatinine were measured, and estimated glomerular filtration rate (eGFR) was calculated. The Kruskal-Wallis H test was used for comparison between groups, and the Spearman test was used for correlation analysis. In CKD stage 3-5, creatinine and iFGF23 levels, as well as the cFGF23/Klotho ratio, were higher (P < 0.01), phosphate levels were higher (P < 0.05), and Klotho levels were lower (P < 0.01), compared with controls. C-terminal-FGF23 levels were higher in CKD phase 4-5 (P < 0.05). In CKD stage 4-5, creatinine, iFGF23, and phosphate levels, as well as the cFGF23/Klotho ratio, were higher (P < 0.01), cFGF23 levels were higher (P < 0.05), and Klotho levels were lower (P < 0.05), compared with CKD stage 3. In CKD stage 5, creatinine and cFGF23 levels, as well as the cFGF23/Klotho ratio, were higher (P < 0.01), phosphate and iFGF23 levels were higher (P < 0.05), and Klotho levels were lower (P < 0.01), compared with CKD stage 4. Phosphate was positively correlated with the cFGF23/Klotho ratio (r = 0.235, P < 0.01). EGFR reduction was associated with an increased cFGF23/Klotho ratio, and the cFGF23/Klotho ratio was positively correlated with phosphate. This suggests that the phosphate level can be controlled by modifying the cFGF23/Klotho ratio.