32P-postlabeling analysis of DNA adduction in mouse skin following topical administration of (+)-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene.
32P-Postlabeling was employed for analysis of DNA adducts produced in mouse skin following topical administration of enantiomers of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol). Deoxynucleoside 3'-monophosphates were isolated by digestion of epidermal DNA with micrococcal endonuclease and spleen phosphodiesterase and phosphorylated with [gamma-32P]ATP. 32P-Labeled deoxynucleoside 3',5'-bisphosphate adducts to diastereomeric benzo[a]pyrene dihydrodiol epoxides (BPDE) were separated by four-directional thin-layer chromatography on poly(ethylenimine)-cellulose plates using a recently described solvent system [Reddy, A. P., Pruess-Schwartz, D., and Marnett, L. J. (1992) Chem. Res. Toxicol. (preceding paper in this issue)]. When (+)-BP-7,8-diol was topically administered, a major adduct spot was detected that cochromatographed with a standard produced by reaction of 7(S),8(R)-dihydroxy-9-(S),10(R)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-syn-BPDE] with DNA. The level of this adduct increased in a dose- and time-dependent fashion and was elevated in animals pretreated with beta-naphthoflavone. Relatively small amounts of radioactivity cochromatographed with standards of deoxynucleoside 3',5'-bisphosphate adducts derived from 7(S),8(R)-dihydroxy-9(R),10(S)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(-)-anti-BPDE]. Following topical administration of (-)-BP-7,8-diol, a single adduct spot was detected that cochromatographed with a standard of the major deoxyguanosine adduct derived from 7(R),8(S)-dihydroxy-9-(S),10(R)-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-anti-BPDE]. The stereochemistry of epoxidation of the enantiomers of BP-7,8-diol indicates that cytochrome P-450 catalyzes the terminal activation step of benzo[a]pyrene activation to an ultimate carcinogen in mouse skin, a target organ for its carcinogenic activity.
Reddy
A P
AP
Department of Chemistry, Wayne State University, Detroit, Michigan 48202.
Pruess-Schwartz
D
D
Ji
C
C
Gorycki
P
P
Marnett
L J
LJ
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000737:chemistry; D000287:Administration, Topical; D000818:Animals; D002417:Cattle; D002855:Chromatography, Thin Layer; D004247:DNA / Q000737:chemistry / Q000187:drug effects*; D018736:DNA Adducts; D004101:Dihydroxydihydrobenzopyrenes / Q000737:chemistry / Q000633:toxicity*; D005260:Female; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D010545:Peroxides / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012867:Skin / Q000737:chemistry / Q000187:drug effects*; D013950:Thymus Gland / Q000737:chemistry
D018736:DNA Adducts; D004101:Dihydroxydihydrobenzopyrenes; D010545:Peroxides; C043931:benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide-DNA; C017228:benzo(a)pyrene 7,8-dihydrodiol; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D004247:DNA