Hydroxyl Free Radical Adduct of Deoxyguanosine: Sensitive Detection and Mechanisms of Formation |
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| Authors: | Robert A Floyd Julia J Watson Peter K Wong Dale H Altmiller Robert C Rickard |
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| Institution: |
a Oklahoma Medical Research Foundation, Oklahoma Children's Memorial Hospital, Oklahoma City, OK, USA
b Departments of Biochemistry and Molecular Biology Oklahoma Children's Memorial Hospital, Oklahoma City, OK, USA
c Department of Pathology, University of Oklahoma Health Sciences Center, Oklahoma Children's Memorial Hospital, Oklahoma City, OK, USA
d Oklahoma Children's Memorial Hospital, Oklahoma City, OK, USA |
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| Abstract: | DNA or 2-deoxyguanosine reacts with hydroxyl free radical to form 8-hydroxy-deoxyguanosine (8-OH-dG). We found that 8-OH-dG can be effectively separated from deoxyguanosine by high pressure liquid chromatography and very sensitively detected using electrochemical detection. The sensitivity by electrochemical detection is about one-thousand fold enhanced over optical detection. Utilizing deoxyguanosine in bicarbonate buffer it was found that ferrous ion, but not ferric ion, was effective in forming 8-OH-dG. The hydroxyl free radical scavenging agents, thiourea and ethanol, were very effective in quenching Fe(11) mediated 8-OH-dG formation, but superoxide dismutase had very little effect. |
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| Keywords: | oxygen free radical hydroxyl free radical adduct DNA damage 2-deoxyguanosine hydroxylation HPLC electrochemical detection hydroxyl radical generation system |
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