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Anal Chem. 2013 Mar 19;85(6):3190-7. doi: 10.1021/ac3034695. Epub 2013 Mar 8.

An ammonium bicarbonate-enhanced stable isotope dilution UHPLC-MS/MS method for sensitive and accurate quantification of acrolein-DNA adducts in human leukocytes.

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State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, China.


Acrolein (Acr), a ubiquitous environmental pollutant, can react directly with genomic DNA to form mutagenic adducts without undergoing metabolic activation. To sensitively and accurately quantify Acr-DNA adducts (including structural isomers and stereoisomers) in human leukocytes, we developed an enhanced stable isotope dilution ultrahigh performance liquid chromatography (UHPLC)-tandem mass spectrometry (MS/MS) method using ammonium bicarbonate (NH4HCO3), which is thermally unstable and degrades readily to carbon dioxide and ammonia in heated gas phase. Interestingly, ammonium bicarbonate (as an additive to the mobile phase) not only improves the protonation of AcrdG adducts but also suppresses the formation of MS signal-deteriorating metal-AcrdG complexes during electrospray ionization, leading to the enhancement of their MS detection by 2.3-8.7 times. In contrast, routinely used ammonium salts (ammonium acetate and ammonium formate) and formic acid do not show similar enhancement. The developed method is potentially useful for enhancing ESI-MS detection of other modified 2'-deoxyribonucleosides that have difficulty in protonation and may form excess metal complexes during electrospray ionization. The limits of detection (LODs, S/N = 3) are estimated to be about 40-80 amol. By the use of the developed method, we found that the Acr adducts of three nucleotides (dG, dA, and dC) can be detected in human leukocytes. In addition to the known γ-AcrdG, α-AcrdA is also identified as an Acr-adduct of high abundance (2.5-20 adducts per10(8) nts).

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