Aflatoxin-guanine DNA Adducts and Oxidatively-induced DNA Damage in Aflatoxin-treated Mice in vivo as Measured by Liquid Chromatography-Tandem Mass Spectrometry with Isotope-dilution.

Dietary exposure to aflatoxin B-1 (AFB(1)) is a significant contributor to the incidence of hepatocellular carcinomas globally. AFB, exposure leads to the formation of AFB(1)-N-7-guanine (AFB(1)-N-7-Gua) and two diastereomers of the imidazole ring-opened 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl-formamido)-9-hydroxyaflatoxin B-1 (AFB(1)-FapyGua) in DNA. These adducts lead to G -> T transversion mutations with the ring-opened adduct being more mutagenic than the cationic species. Accurate measurement of these three adducts as biomarkers in DNA and urine will help identify dietary exposure to AFB(1) as a risk factor in the development of hepatocellular carcinoma worldwide. Herein, we report an improved methodology for the measurement of AFB(1)-N-7-Gua and the two diastereomers of AFB(1)-FapyGua using liquid chromatography-tandem mass spectrometry with isotope dilution. We measured the levels of these compounds in liver DNA of six control mice and six AFB(1)-treated mice. Levels varying from 1.5 to 45 lesions/10(6) DNA bases in AFB(1)-treated mice were detected depending on the compound and animal. No background levels of these adducts were detected in control mice. We also tested whether the AFB(1) treatment caused oxidatively induced DNA base damage using gas chromatography-tandem mass spectrometry with isotope dilution. Although background levels of several pyrimidine- and purine-derived lesions were detected, no increases in these levels were found upon AFB(1) treatment of mice. On the other hand, significantly increased levels of (5'R)- and (5'S)-8,5'-cyclo-2'-deoxyadenosines were observed in liver DNA of AFB(1)-treated mice. The impact of this work is expected to achieve the accurate measurement of three AFB(1)-DNA adducts and oxidatively induced DNA lesions as biomarkers of AFB(1) exposure as germane to investigations designed for the prevention of aflatoxin-related hepatocellular carcinomas and for determining the effects of genetic deficiencies in human populations.