Suppression of adventitious formation of 8-oxoguanine(TMS)4 from guanine during trimethylsilylation.
GC/MS quantitation of 8-oxoguanine derived from DNA may yield artificially high values when trimethylsilylation is used as the technique to form a volatile derivative. Significant quantities of the tetrakis trimethylsilyl-derivative of 8-oxoguanine may be formed from guanine during the derivatization reaction at elevated temperatures. We have screened eight antioxidants in an attempt to identify a substance that will reduce the spurious formation of this product. Whereas several compounds were active, N-phenyl-1-naphthylamine was the most effective agent. Moreover, this compound was sufficiently basic to act as a catalyst for the derivatization reaction. Our data, which are based on the use of an isotopically labeled internal standard, show that the level of the tetrakis trimethylsilyl derivative of 8-oxoguanine remained constant as a function of derivatization heating time when N-phenyl-1-naphthylamine was added to a reaction in which guanine was present. In contrast, the level in control samples increased linearly with reaction time indicating formation of 8-oxoGua during derivatization. The level of 8-oxoguanine was measured in both a synthetic oligodeoxynucleotide containing a single 8-oxo-2'-deoxyguanosine moiety and commercially available calf thymus DNA using this technique.
Hong
J
J
Department of Pharmacological Sciences, State University of New York at Stony Brook, Stony Brook, NY 11794-3400, USA.
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Johnson
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D015057:1-Naphthylamine / Q000031:analogs & derivatives*; D000818:Animals; D000975:Antioxidants; D001483:Base Sequence; D002417:Cattle; D002623:Chemistry Techniques, Analytical; D004247:DNA / Q000737:chemistry; D008401:Gas Chromatography-Mass Spectrometry; D006147:Guanine / Q000031:analogs & derivatives* / Q000138:chemical synthesis* / Q000737:chemistry*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D013696:Temperature; D014297:Trimethylsilyl Compounds / Q000138:chemical synthesis* / Q000737:chemistry
D000975:Antioxidants; D009838:Oligodeoxyribonucleotides; D014297:Trimethylsilyl Compounds; C024829:8-hydroxyguanine; D006147:Guanine; C005444:N-phenyl-1-naphthylamine; D004247:DNA; D015057:1-Naphthylamine