Characterization of radiation-induced products of thymidine 3'-monophosphate and thymidylyl (3'-->5') thymidine by high-performance liquid chromatography and laser-desorption Fourier-transform mass spectrometry.
High-performance liquid chromatography (HPLC) and laser-desorption Fourier-transform mass spectrometry (LD FTMS) have been applied for direct measurements of radiation-induced products of nucleic acid constituents containing thymidine. Laser desorption FTMS could be used for the direct detection (neither hydrolyzed nor derivatized) of X ray-induced decomposition products of aqueous thymidine monophosphate. After these initial experiments, a variety of hydrogenated and hydroxylated thymine standards were acquired and examined by FTMS to assist in the identification of unknown radiation-induced decomposition products of thymine-containing nucleotides and dinucleotides. To extend these studies to dinucleotides, the radiation-induced products generated by the gamma radiolysis of thymidylyl (3'-->5') thymidine (TpT) were isolated by reverse-phase HPLC and identified by LD FTMS. Thymine and thymidine 3'-monophosphate were observed as the major products in this case. Several of the minor products of the HPLC profile were pooled in a single fraction and characterized simultaneously by LD FTMS. The resulting mass spectra indicated the presence of hydroxy-5,6-dihydrothymidine monophosphate, 5,6-dihydrothymidine monophosphate and thymidine monophosphate, thymine glycol, hydroxy-5,6-dihydrothymine, 5-hydroxy-methyluracil and 5,6-dihydrothymine. The combination of HPLC purification and LD FTMS structural characterization provides a useful tool for the direct measurement of radiation-induced products of nucleotides and dinucleotides.
Yoshida
H
H
Oak Ridge National Laboratory, Tennessee 37831-6120.
Hettich
R L
RL
D002851:Chromatography, High Pressure Liquid; D005583:Fourier Analysis; D013058:Mass Spectrometry; D009841:Oligonucleotides / Q000737:chemistry / Q000528:radiation effects; D013942:Thymine Nucleotides / Q000737:chemistry / Q000528:radiation effects*; D014965:X-Rays
D009841:Oligonucleotides; D013942:Thymine Nucleotides