Technique for detection of DNA nucleobases by reversed-phase high-performance liquid chromatography optimized for...

A reversed-phase high-performance liquid chromatographic (HPLC) method for quantification of iododeoxyuridine (IdUrd) substitution for thymidine in DNA was developed. IdUrd substitution was determined by HPLC with ultraviolet absorbance in two cell lines (L1210 and HT-29) after incubation in vitro with IdUrd alone or with IdUrd and fluorodeoxyuridine. In addition, radiolabeled IdUrd was used concomitantly with HPLC to evaluate the degree of dehalogenation. This HPLC technique has also been applied to the measurement of IdUrd incorporation in vivo into DNA from peripheral granulocytes of patients receiving IdUrd. This method provides an interesting tool for the quantification of drug substitution into DNA and can be applied to multiple sampling sites in animal and human studies.