2--deoxyguanosine-5--phosphate and styrene-oxide

Lamination workers are exposed to large amounts of styrene. A postlabelling method was developed in order to detect DNA adducts in white blood cells from such workers. First, styrene oxide was reacted with DNA, and the adducts were characterized by the nuclease P1 version of the 32P-postlabelling technique. The adducts in human samples were transferred magnetically during chromatography. The 2'-deoxyguanosine 3'-monophosphate (dGMP) adduct standards, including N2 and O6 adducts, were shown to be resistant to the action of nuclease P1. The O6 adducts were detected in the femtomole range at about 10% labelling efficiency. In lamination workers, the level of O6 adducts, adjusted for adduct recovery, was 5/10(8) nucleotides, which was five times the level in controls.

Topics: Autoradiography; Chromatography, Thin Layer; Deoxyguanine Nucleotides; DNA; Epoxy Compounds; Humans; Occupational Exposure; Phosphorus Radioisotopes; Single-Strand Specific DNA and RNA Endonucleases

Adducts were prepared by reacting styrene oxide with 2-deoxyguanosine 3'-monophosphate (dGMP). Four isomeric N-7-, two diastereomeric N2- and three isomeric O6-adduct were isolated and characterized. The adducts were used as substrates in the 32P-postlabeling reaction. No phosphorylation products were seen with the N-7-alkylation products. One diastereomeric N2-adduct was labeled with 20% efficiency and the second with a markedly lower efficiency. Two of the three O6-adducts were labeled with 5% and the third with 10% labeling efficiency. The results suggest that large N-7-dGMP adducts are very poor substrates of T4 polynucleotide kinase. The diastereomeric products are labeled at different efficiencies indicating stereoselectivity in the kinase reaction.

Topics: Deoxyguanine Nucleotides; Epoxy Compounds; Phosphorus Radioisotopes; Phosphorylation; Polynucleotide 5'-Hydroxyl-Kinase; Stereoisomerism

Deoxyguanosine 3'-monophosphate (dGMP) was alkylated at the 7-position by dimethyl sulfate, ethylene oxide and styrene oxide in aqueous media and glacial acetic acid, respectively, to yield reasonable quantities of the products, which were purified by HPLC. dGMP adducts are needed as standards for the 32P-postlabelling assay. The stability of the adducts was studied at 37 degrees and neutral pH. The half-lives of disappearance of 7-methyl-dGMP and the beta-isomers of the styrene oxide adducts were about 250 min; 7-hydroxy-ethyl-dGMP and the alpha-isomers of the styrene oxide adducts had respective half-lives of 340 and 440 min. In all cases the main degradation product was the corresponding guanine adduct. The results demonstrate considerable lability of the 7-alkylation products of dGMP which has to be taken into consideration in devising the 32P-postlabelling assay.

Topics: Alkylating Agents; Chromatography, High Pressure Liquid; Deoxyguanine Nucleotides; Epoxy Compounds; Ethers, Cyclic; Ethylene Oxide; Guanine Nucleotides; Guanosine Diphosphate; Methylation; Sulfuric Acid Esters; Sulfuric Acids

The 32P-postlabelling procedure has been used to detect styrene oxide (SO)-DNA adducts. Reactions of SO with DNA and dGMP in vitro produced adducts that were similar, indicating that dGMP was the primary base for modification in DNA. Two SO adducts were also detected in DNA isolated from lymphocytes of a styrene-exposed worker but not in DNA from an unexposed worker. These results indicate that 32P-postlabelling can be used for quantification of DNA adducts in workers exposed to styrene.

Topics: Deoxyguanine Nucleotides; DNA; Environmental Exposure; Environmental Monitoring; Epoxy Compounds; Ethers, Cyclic; Humans; Lymphocytes; Phosphorus Radioisotopes