Introduction: In patients with chronic kidney disease (CKD), uremic toxins accumulate in blood and cannot be excreted with urine. Accumulation of these toxins has negative effects on many body functions. Because of the importance of these toxins, we developed and validated a simple, sensitive, accurate, and precise method for the determination of two main uremic toxins:phenol and p-cresol in human urine. Materials and methods:Separation of these analytes in urine samples was achieved by reverse-phase high-performance liquid chromatography (RP-HPLC) with a C18 column at 35 degrees C using the mobile phase of methanol-water (65:35) at a flow rate of 1.4 mL min(-1). Fluorimetric detection was used at 284 nm for excitation and 310 nm for emission. Results:The method is linear over the range of 1.5-35 ng mL(-1) and 1-45 ng mL(-1) for phenol and p-cresol, respectively. The method was applied to urine samples from 10 healthy subjects and 10 chronic kidney disease patients. Conclusions:This assay appears to be useful in routine analysis of clinical samples for simultaneous determination of phenol and p-cresol levels in urine.