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Updated: Jul 9, 2026
Formation of Covalent DNA Adducts by Enzymatically Activated Carcinogens and Drugs In Vitro and Their Determination by 32P-postlabeling
Published on: March 20, 2018
Detection of the acrolein-derived cyclic DNA adduct by a quantitative 32P-postlabeling/solid-phase extraction/HPLC
Armaghan Emami1, Marcin Dyba, Amrita K Cheema
1Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA.
Abstract:
Acrolein (Acr), a hazardous air pollutant, reacts readily with deoxyguanosine (dG) in DNA to produce cyclic 1, N2-propanodeoxyguanosine adducts (Acr-dG). Studies demonstrate that these adducts are detected in vivo and may play a role in mutagenesis and carcinogenesis. In the study described here, a quantitative 32P-postlabeling/solid-phase extraction/HPLC method was developed by optimizing the solid-phase extraction and the 32P-postlabeling conditions for analysis of Acr-dG in DNA samples with a detection limit of 0.1 fmol. It was found that Acr-dG can form as an artifact during the assay. Evidence obtained from mass spectrometry indicates that the Acr in water used in the assay is a likely source of artifact formation of Acr-dG. The formation of Acr-dG as an artifact can be effectively blocked by adding glutathione (GSH) to the DNA sample to be analyzed. In addition, Acr-dG was detected as a contaminant in the commercial dG and dT 3'-monophosphate samples. Finally, this method was used to detect Acr-dG in calf thymus and human colon HT29 cell DNA with an excellent linear quantitative relationship.
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