Development and Application of an LC-MS/MS Method for the Detection of the Vinyl Chloride-Induced DNA Adduct N2,3-Ethenoguanine in Tissues of Adult and Weanling Rats Following Exposure to [13C2]-VC

In the 1970s, exposure to vinyl chloride (VC) was shown to cause liver angiosarcoma in VC workers, We have developed a new LC-MS/MS method for analyzing the promutagenic DNA adduct N-2,3-ethenoguanine (epsilon G) and have applied this to DNA from tissues of both adult and weanling rats exposed to 1100 ppm [C-13(2)]-VC for 5 days or 1100 ppm VC for I day. This assay utilizes neutral thermal hydrolysis and an HPLC cleanup prior to quantitation by LC-MS/MS. The number of endogenous and exogenous EG adducts in DNA from tissues of adult rats exposed to [C-13(2)]-VC for 5 days was 4.1 +/- 2.8 adducts/10(8) guanine of endogenous and 19.0 +/- 4.9 adducts/108 guanine of exogenous EG in the liver, 8.4 +/- 2.8 adducts/10(8) guanine of endogenous and 7.4 +/- 0.5 adducts/108 guanine of exogenous epsilon G in the lung, and 5.9 +/- 3.3 adducts/10(8) guanine of endogenous and 5.7 +/- 2.1 adducts/10(8) guanine of exogenous epsilon G in the kidney (a = 4). Additionally, the data from weanling rats demonstrated higher numbers of exogenous epsilon G, with 4-fold higher amounts in the liver DNA of weanlings (75.9 +/- 17.9 adducts/108 guanine) in comparison to adult rats and similar to 2-fold higher amounts in the lung (15.8 +/- 3.6 adducts/10(8) guanine) and kidney (12.9 +/- 0.4 adducts/108 guanine) (a = 8). The use of stable isotope labeled VC permitted accurate estimates of the half-life of epsilon G for the first time by comparing [C-13(2)]-epsilon G in adult rats with identically exposed animals euthanized 2, 4, or 8 weeks later. The half-life of epsilon G was found to be 150 days in the liver and lung and 75 days in the kidney, suggesting little or no active repair of this promutagenic adduct.