Urinary and serum pseudouridine concentrations were determined by high-performance liquid chromatography in 80 patients with primary liver cancer, 32 with benign space occupying lesions of the liver, 42 with liver cirrhosis and 40 healthy subjects. Their mean urinary and serum pseudouridine levels were 39.2 +/- 11.5 nmol/mumol creatinine and 3.4 +/- 1.3 mumol/L, 24.5 +/- 5.4 nmol/mumol creatinine and 2.5 +/- 0.5 mumol/L, 22.8 +/- 7.8 nmol/mumol creatinine and 2.3 +/- 0.4 mumol/L, 26.4 +/- 4.6 nmol/mumol creatinine and 2.3 +/- 0.4 mumol/L, respectively. Exceeding the mean plus 2SD of pseudouridine of healthy control was considered as positive value for the diagnosis of primary liver cancer. Thus the positivity of urinary and serum pseudouridine in hepatoma was 71.3% and 70.0%, respectively. The positive rate of combined pseudouridine and alpha-fetoprotein assay was 91.3% in patients with hepatoma. Besides, pseudouridine levels could elevate before positive localization and reduce to normal levels after tumor resection. The results showed that the determination of pseudouridine is of clinical significance in the diagnosis and monitoring of primary liver cancer.