Preparation and characterization of carbon-13, nitrogen-15 labeled nucleosides, oxidized nucleosides and DNA

Oxidative damage to DNA has been implicated in the etiology of various human diseases. We are developing strategies to test the hypothesis that progressive diseases of aging, such as Alzheimer's are the product of accumulated and unrepaired DNA damage to neurons. These strategies focus on the direct measurement of oxidized nucleosides in human brain tissue DNA. One target analyte is thymidine glycol. Stable isotope labeled internal standards for the quantification of the major nucleosides have not been commercially available for research purposes. The recent availability of highly enriched $\sp{13}$C, $\sp{15}$N algal DNA has provided a new source for labeled double stranded DNA, for labeled nucleosides and for chemically modified nucleosides to use as internal standards. To utilize algal DNA, we have tested methods of DNA enzymatic cleavage, chromatographic separation of the nucleosides, and chemical methods of oxidation suitable for preparing $\mu$g to mg quantities of well characterized material using labeled nucleotides. The characterization of $\sp{13}$C, $\sp{15}$N nucleosides and thymidine glycol by NMR and mass spectra indicate that these compounds have been successfully isolated and purified.