Analysis of 4‑Hydroxy-1-(3-pyridyl)-1-butanone (HPB)-Releasing DNA Adducts in Human Exfoliated Oral Mucosa Cells by Liquid Chromatography–Electrospray Ionization–Tandem Mass Spectrometry

• Published in: Chemical research in toxicology 2013-01, Vol.26 (1), p.37-45
• Main Authors: Stepanov, Irina, Muzic, John, Le, Chap T, Sebero, Erin, Villalta, Peter, Ma, Bin, Jensen, Joni, Hatsukami, Dorothy, Hecht, Stephen S
• Format: Article
• Language: English
• Subjects: Biomarkers - urine; Butanones - chemistry; Butanones - toxicity; Chromatography, High Pressure Liquid; DNA - chemistry; DNA - metabolism; DNA Adducts - analysis; DNA Adducts - isolation & purification; Humans; Mouth Mucosa - chemistry; Mouth Mucosa - drug effects; Mouth Mucosa - metabolism; Mouthwashes - chemistry; Nitrosamines - chemistry; Nitrosamines - metabolism; Pyridines - chemistry; Pyridines - toxicity; Smoking; Solid Phase Extraction; Spectrometry, Mass, Electrospray Ionization
• ISSN: 0893-228X; 1520-5010; 1520-5010
• DOI: 10.1021/tx300282k

Quantitation of DNA adducts could provide critical information on the relationship between exposure to tobacco smoke and cancer risk in smokers. In this study, we developed a robust and sensitive liquid chromatography–tandem mass spectrometry method for the analysis of 4-hydroxy-1-(3-pyridyl)-1-butanone (HPB)-releasing DNA adducts in human oral cells, a noninvasive source of DNA for biomarker studies. Isolated DNA undergoes acid hydrolysis, after which samples are purified by solid-phase extraction and analyzed by LC-ESI-MS/MS. The developed method was applied to the analysis of samples obtained via collection with a commercial mouthwash from 30 smokers and 15 nonsmokers. In smokers, the levels of HPB-releasing DNA adducts averaged 12.0 pmol HPB/mg DNA (detected in 20 out of 28 samples with quantifiable DNA yield), and in nonsmokers, the levels of adducts averaged 0.23 pmol/mg DNA (detected in 3 out of 15 samples). For the 30 smoking subjects, matching buccal brushings were also analyzed, and HPB-releasing DNA adducts were detected in 24 out of 27 samples with quantifiable DNA yield, averaging 44.7 pmol HPB/mg DNA. The levels of adducts in buccal brushings correlated with those in mouthwash samples of smokers (R = 0.73, p < 0.0001). Potentially, the method can be applied in studies of individual susceptibility to tobacco-induced cancers in humans.