| Summary: | 碩士 === 國立中正大學 === 化學研究所 === 89 === Etheno DNA adducts have been detected in tissue DNA of rodents and humans from various exogenous and endogenous sources. In this report, three assays were developed for detection and quantification of 3,N4-ethenocytosine (eCyt), namely by HPLC/fluorescence and gas chromatography/negative chemical ionization mass spectrometry (GC/NCI/MS). Direct measurement of 3,N4-etheno-2'-deoxycytidine (edCyd) gave the limit of detection (LOD) of 4.0 pmol, and the limit of quantification (LOQ) of 5.9 pmol, which was lower than the published method. The other HPLC/fluorescence assay was developed for the non-fluorescent eCyt after derivatization with 4-bromomethyl-6,7-dimethoxycoumarin (Br-MDC). The LOD for MDC-eCyt is 34 fmol, and the LOQ is 6.5 pmol. An isotope dilution GC/NCI/MS assay was developed for detection and quantification of eCyt in DNA using [13C4, 15N3]eCyt as the internal standard. The adduct-enriched fraction of DNA hydrolysate was derivatized with pentafluorobenzyl (PFB) bromide before GC/NCI/MS analysis. The LOD for PFB-eCyt was 1.0 fg (3.2 amol, S/N = 41) injected on column, and the LOQ was 1.0 pg (7.4 fmol) of eCyt, which was about one thousand times lower than that of the HPLC/fluorescence assay for the edCyd. Analysis of chloroacetaldehyde-treated calf thymus DNA by this GC/NCI/MS assay and by HPLC/fluorescence gave similar adduct levels. This GC/NCI/MS assay was used for analysis of eCyt and edCyd in human urine and the levels for a smoker were 107 and 186 pg/ml, respectively.
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