/sup 32/P-Postlabeling test for covalent DNA binding of chemicals in vivo: Application to a variety of aromatic carcinogens and methylating agents

Journal Article · Wed Feb 01 04:00:00 EST 1984 · Carcinogenesis (N.Y.); (United States)

DOI:https://doi.org/10.1093/carcin/5.2.231· OSTI ID:6850981

Reddy, M V; Gupta, R C; Randerath, E; Randerath, K

Carcinogen--DNA adducts were detected and determined by /sup 32/P-postlabeling assay after exposure of mouse or rat tissues in vivo to a total of 28 compounds comprising 7 arylamines and derivatives, 3 azo compounds, 2 nitroaromatics, 12 polycyclic aromatic hydrocarbons, and 4 methylating agents. DNA was isolated from mouse skin, mouse liver, and rat liver after treatment with the individual carcinogens, then digested enzymatically to deoxyribonucleoside 3'-monophosphates, which were converted to 5'-/sup 32/P-labeled deoxyribonucleoside 3',5'-bisphosphates by T4 polynucleotide kinase-catalyzed (/sup 32/P)phosphate transfer from (gamma-/sup 32/P)ATP. The nucleotides were resolved by anion-exchange t.l.c. on polyethyleneimine-cellulose and detected by autoradiography. The determination of low levels of DNA binding of the aromatic carcinogens entailed the removal of normal nucleotides prior to the resolution of adduct nucleotides. For this purpose, an alternative procedure employing reversed-phase t.l.c. was devised which offered advantages for the detection of quantitatively minor adducts. The procedures described enabled the detection of 1 aromatic DNA adduct in approximately 10(/sup 8/) normal nucleotides, while the limit of detection of methylated adducts was 1 adduct in approximately 6 X 10(/sup 5/) nucleotides. The results show that a great number of carcinogen-DNA adducts of diverse structure are substrates for /sup 32/P-labeling by polynucleotide kinase-catalyzed phosphorylation. Because covalent DNA adduct formation in vivo appears to be an essential property of the majority of chemical carcinogens, /sup 32/P-postlabeling analysis of carcinogen--DNA adducts in mammalian tissues may serve as a test for the screening of chemicals for potential carcinogenicity.

• Research Organization: Department of Pharmacology, Baylor College of Medicine, Houston, TX
• OSTI ID: 6850981
• Journal Information: Carcinogenesis (N.Y.); (United States), Journal Name: Carcinogenesis (N.Y.); (United States) Vol. 5:2; ISSN CRNGD
• Country of Publication: United States
• Language: English

Related Subjects

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560301 -- Chemicals Metabolism & Toxicology-- Cells-- (-1987)
59 BASIC BIOLOGICAL SCIENCES
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ADDUCTS
ALKYLATING AGENTS
AMINES
ANIMALS
AROMATICS
ATP
AUTORADIOGRAPHY
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL EFFECTS
CARCINOGEN SCREENING
CELL CULTURES
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DNA ADDUCTS
HYDROCARBONS
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MAMMALS
MICE
NITRO COMPOUNDS
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
POLYCYCLIC AROMATIC AMINES
POLYCYCLIC AROMATIC HYDROCARBONS
POLYCYCLIC NITRO COMPOUNDS
RADIOISOTOPES
RATS
RODENTS
SCREENING
SEPARATION PROCESSES
STRUCTURE-ACTIVITY RELATIONSHIPS
THIN-LAYER CHROMATOGRAPHY
TRACER TECHNIQUES
VERTEBRATES