Simultaneous detection of five different 2-hydroxyethyl-DNA adducts formed by ethylene oxide exposure, using a high-performance liquid chromatography/electrospray ionisation tandem mass spectrometry assay.
A method has been developed for the simultaneous detection and quantitation of five different 2-hydroxyethyl-DNA (HE-DNA) adducts that could be formed as a result of exposure to ethylene oxide (EO). In addition to the major N7-HE-guanine (N7-HEG) adducts this assay can also measure the less prevalent but potentially more biologically significant N1-HE-2'-deoxyadenosine (N1-HEdA), O(6)-HE-2'-deoxyguanosine (O(6)-HEdG), N(6)-HE-2'-deoxyadenosine (N(6)-HEdA) and N3-HE-2'-deoxyuridine adducts (N3-HEdU). The method involves the isolation of HE adducts from the unmodified nucleosides by either neutral thermal hydrolysis or enzymatic digestion, followed by high-performance liquid chromatographic (HPLC) purification, before detection and quantification by liquid chromatography tandem mass spectrometry (LC/MS/MS) using selective reaction monitoring (SRM). The limits of detection were in the range 0.5-25 fmol for each individual adduct, making this one of the most sensitive assays available for the detection of N7-HEG. To illustrate the possible applications of the assay, it has been employed in the measurement of endogenous/background and EO-induced HE adducts in a variety of DNA samples.
Cancer Biomarkers and Prevention Group, Department of Cancer Studies and Molecular Medicine, The Biocentre, University of Leicester, University Road, Leicester LE1 7RH, UK.
Elaine M
EM
Tompkins
Donald J L
DJ
Jones
John H
JH
Lamb
Debbie A
DA
Marsden
Peter B
PB
Farmer
Karen
K
Brown
D000818:Animals; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D018736:DNA Adducts / Q000032:analysis* / Q000138:chemical synthesis; D005027:Ethylene Oxide / Q000737:chemistry*; D008099:Liver / Q000737:chemistry; D051381:Rats; D011916:Rats, Inbred F344; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D013056:Spectrophotometry, Ultraviolet; D053719:Tandem Mass Spectrometry
D018736:DNA Adducts; D005027:Ethylene Oxide