Deoxynucleotide modifications

Steinberg, J. J., and Cajigas, A. (1992). Enzymatic shot-gun 5 -phosphorylation and 3 -sister phosphate exchange a two-dimensional thin-layer chromatographic technique to measure DNA deoxynucleotide modification. J. Chromatogr. 574 41-55. 

Specific DNA adducts are excreted into urine as modified bases and deoxynucleotides [121-123]. As endonucleases and glycosylases repair the damaged DNA, deoxynucleotides and bases respectively are liberated. Oligonucleotides and bulky DNA adducts like benzo[a]pyrene-deoxynucleotides or aflatoxin-Bj-Ny-dG are also excreted. Immuno-affinity chromatography, GC/MS, LC-MS/MS,or HPLE-EC can be used to quantify the wide range of base modifications and thus to estimate the DNA repair activity, whereas ELISA assays do not yet have sufficient proven specificity. 

Cyclic nucleotides under our conditions have a long retention time, and are reasonably separated from the ribonucleotides and deoxynucleotides (c-CMP at 5.9, c-GMP at 36.4, and c-AMP at 112 minutes). However the time of elution of cAMP is longer than normally desired with an HPLC system. It is evident from Table 1 and Fig. 3, that modification of the eluant by the addition of differing amounts of methanol, expedites the elution without affecting resolution. From Fig. 3 it is also clear that the retention time of CTP, GTP, and ATP is influenced by the incorporation of methanol in the eluant. 

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