INTERFERENCE OF A METHOTREXATE DERIVATIVE WITH URINARY ONCOPTERIN [N-2-(3-AMINOPROPYL)BIOPTERIN] MEASUREMENT BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH FLUOROMETRIC DETECTION

We previously reported a HPLC assay method using fluorimetric detectio n for the simultaneous determination of urinary N-2-(3-aminopropyl)bio pterin (oncopterin, a natural pteridine newly found in urine from canc er patients), biopterin and neopterin. We now have observed that an un known substance, which may be derived from methotrexate, in urine from a patient with stomach cancer interfered with the assay of oncopterin and demonstrated that oncopterin could be completely separated from t he unidentified substance by HPLC using a Nucleosil 100-5SA strong cat ion-exchange column. Furthermore, oncopterin was not detectable by thi s HPLC-fluorimetric method in urine samples from patients with stomach cancer who were not treated with methotrexate. The content of urinary oncopterin from cancer patients is supposed to be very low, with less than 1 mu mol/mol creatinine. The present results indicate that the p eak found with elution from the C-18 column was a methotrexate-derived compound and co-eluted with the analyte oncopterin.