Liquid chromatographic determination of urinary 5-methyl-2 '-deoxycytidineand pseudouridine as potential biological markers for leukaemia

A simple reversed-phase liquid chromatographic (LC) method for the determin ation of urinary 5-methyl-2'-deoxy-cytidine (m(5)dCyd), recently claimed (o n the basis of an imuno-technique) to be a potential marker for leukaemia, has been developed. Sample pre-treatment is based on a microcolumn clean-up step with an average recovery of 79% and a RSD of 3%. Detection limit was 0.2 mu g/ml which is about tenfold lower than levels previously measured by an ELISA method in urine of healthy individuals. The creatinine (Cre) excr etion, necessary for normalising the m(5)dCyd excretion, was evaluated by i on-pair liquid chromatography which permitted the simultaneous determinatio n of pseudouridine (Psi), a modified nucleoside also potentially useful as a marker for leukaemia. The described LC procedures were applied to the ana lysis of urine samples from healthy individuals and leukaemia patients. Whi le the urinary Psi/Cre ratio was found significantly increased for leukaemi a patients, the urinary m(5)dCyd levels in healthy individuals were below t he detection limits and did not increase in presence of the malignant disea se. (C) 1999 Elsevier Science B.V. All rights reserved.