Inhibition of clastogenicity of benzo[a]pyrene and of its trans-7,8-dihydrodiol in mice in vivo by fruits, vegetables, and flavonoids.

In the in vivo mouse bone marrow micronucleus assay, homogenates of spinach, artichoke, peaches, and blue grapes as well as commercial concentrates of these vegetables and fruits reduced induction of micronuclei by benzo[a]pyrene (BaP) by 43-50%. Concentrates of strawberries (31% reduction) and of cauliflower (20% reduction) were less potent. Inhibition of genotoxicity by spinach and peaches was not caused by any delay in maturation of micronucleated erythrocytes as shown by experiments with sampling times of 24, 48, and 72 h after dosing of BaP. Pre-treatment of the mice with spinach 48, 24, and 12h before application of BaP resulted in a 44% reduction of micronuclei while peaches generated only a marginal effect. A post-treatment procedure administering spinach or peaches 6h after dosing of BaP did not indicate any protective effects. When trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BaP-7,8-OH) was applied for induction of micronuclei spinach and peaches reduced the number of micronuclei by 55 and 48%, respectively. Pre-treatment of mice with spinach 96, 72, and 60 h before sacrifice caused a decline of hepatic 7-ethoxyresorufin-O-dealkylase (EROD) and of 7-pentoxyresorufin-O-dealkylase (PROD) activities by factors of 2.2 and 1.4, respectively. However, statistical significance was not reached. On the other hand, peaches had no influence on hepatic EROD or PROD activities. The flavonoids quercetin and its glucoside isoquercitrin, administered orally in doses of 0.03 mmol/kg body weight simultaneously with intraperitoneally given BaP, reduced the number of micronuclei in polychromatic erythrocytes of the bone marrow of mice by 73 and 33%. Ten-fold higher concentrations, however, reversed the effects with a particular strong increase observed with isoquercitrin (+109%; quercetin: +16%).

Inflammatory response of mouse skin exposed to the very potent carcinogen dibenzo[a,l]pyrene: a model for tumor promotion.

The potent carcinogenicity of dibenzo[a,l]pyrene in mouse skin is associated with an inflammation unique among polycyclic aromatic hydrocarbons and expressed as erythema. The time course of erythema and the associated histological events in the skin of female SENCAR mice were determined after a single application of 6.25-200 nmol dibenzo[a,l]pyrene or selected metabolites. Dibenzo[a,l]pyrene and dibenzo[a,l]pyrene-11,12-dihydrodiol, precursor to the bay-region diol epoxide, induced an erythema first present 5-6 days after treatment. Dibenzo[a,l]pyrene-8,9-dihydrodiol and other dibenzo[a, l]pyrene metabolites, however, did not induce erythema. These findings suggest a central role for the bay-region diol epoxide in the induction of the observed inflammation. The intensity and duration of erythema were dose-dependent, whereas the delayed appearance of erythema was constant and dose-independent. These results suggest induction of an immune hypersensitivity by dibenzo[a, l]pyrene and its 11,12-dihydrodiol. Histological changes in the skin were consistent with a contact hypersensitivity reaction and included, in association with erythema, epidermal hyperplasia and the presence of mononuclear leukocytes in the dermis. Animals were tested for dibenzo[a,l]pyrene-induced contact hypersensitivity. Female SENCAR mice were treated with a single dermal application of dibenzo[a,l]pyrene or 7,12-dimethylbenz[a]anthracene. Five days later, the animals were challenged with a single application of dibenzo[a,l]pyrene or 7,12-dimethylbenz[a]anthracene to the ear pinna. Ear swelling exhibited features of a contact hypersensitivity reaction, including (1) delayed appearance after challenge, (2) noninducibility in animals not previously exposed to chemical sensitizer, and (3) chemical specificity. The results suggest that dibenzo[a,l]pyrene induces, via its bay-region diol epoxide, a contact hypersensitivity reaction that may promote tumor development and thereby enhance carcinogenic potency.

Southern flounder hepatic and intestinal metabolism and DNA binding of benzo[a]pyrene (BaP) metabolites following dietary administration of low doses of BaP, BaP-7,8-dihydrodiol or a BaP metabolite mixture.

Certain finfish species living in chemically polluted environments exhibit a high incidence of gastrointestinal tract tumors. Carnivorous fish in such environments are likely to consume invertebrates which contain chemical procarcinogens and the invertebrate biotransformation products of these compounds. The retention in tissues, extent of DNA adduct formation in liver and intestine, and metabolite composition of bile was investigated in southern flounder following gavage with pure [3H]- or [14C]benzo[a]pyrene (BaP), pure [14C]benzo[a]pyrene-7,8-dihydrodiol (BaP-7,8D), or hepatopancreas from spiny lobsters previously dosed with [3H]- or [14C]BaP (Metab.HP). Metab.HP contained mainly polar conjugates of BaP diols, triols and tetraols. BaP-7,8D was retained in fish tissues and bile at 24 h to a greater extent (33.6% of the dose), than either BaP (19.00%) or Metab.HP (6.6%). Hepatic and intestinal DNA isolated from all dosed fish contained covalently bound radioactivity, but exposure to BaP-7,8D or BaP resulted in significantly higher binding in both tissues than exposure to Metab.HP. Hepatic DNA from BaP and BaP-7,8D-dosed flounder contained 0.24 +/- 0.07 and 0.33 +/- 0.06 pmol BaP equivalents/mg DNA respectively (mean +/- S.E.), while hepatic DNA isolated from Metab.HP-dosed flounder contained 0.006 +/- 0.002 pmol BaP equivalents/mg DNA. Binding of radioactivity to intestinal DNA was significantly higher than to hepatic DNA for flounder dosed with Metab.HP (0.026 +/- 0.003) or with BaP (0.76 +/- 0.27) but not for flounder dosed with BaP-7,8D (0.44 +/- 0.09). These studies show that dietary BaP, and metabolites likely to be present in invertebrates, can be absorbed by the southern flounder and form DNA adducts in target organs.

Contrasting disposition and metabolism of topically applied benzo(a)pyrene, trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene, and 7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene in mouse epidermis in vivo.

Whereas extensive evidence indicates that 7 beta,8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (anti-BPDE) is a major ultimate carcinogen of benzo(a)pyrene (BaP) in mouse skin, tumorigenicity studies have consistently shown that anti-BPDE is less active then BaP in this model system. In order to investigate factors responsible for this apparent contradiction, we have compared the disposition, metabolism, and DNA binding of [3H]BaP, (+/-)-trans-7,8-[14C]dihydroxy-7,8-dihydrobenzo(a)pyrene [(+/-)-[14C]BaP-7,8-diol), and (+/-)-anti-[3H]BPDE in mouse epidermis in vivo. There were remarkable differences in the total radioactivity recovered in epidermis at various times after topical application of BaP, BaP-7,8-diol, and anti-BPDE. BaP and its metabolites were removed from epidermis gradually (t1/2 approximately equal to 2 h). However, 60-65% of anti-BPDE disappeared from mouse epidermis within 3 min of application, while a second slower phase of removal of radioactivity was observed between 8 min and 2 h. The kinetics of removal of BaP-7,8-diol and its metabolites were intermediate between those of BaP and anti-BPDE. The half-life of anti-BPDE in mouse epidermis was measured by trapping it with 2-mercaptoethanol. The initial half-life was about 6 min, similar to that observed in vitro. However, following the initial rapid penetration of anti-BPDE through epidermis most of the remaining material became immobilized in an epidermal binding site in which its half-life was greater than 2 h. Qualitatively, the metabolite patterns of BaP, BaP-7,8-diol, and anti-BPDE were similar to expectations based on in vitro studies. However, the kinetics of metabolite formation from BaP were different from those of BaP-7,8-diol or anti-BPDE. The extents of formation of anti-BPDE-DNA adducts 24 h after application of BaP, BaP-7,8-diol, or anti-BPDE to mouse skin were similar despite the fact that the levels of anti-BPDE present in epidermis were about 50 to 100 times greater after application of BaP-7,8-diol or anti-BPDE than after application of BaP. The results of this study demonstrate that the quantitative aspects of BaP-7,8-diol and anti-BPDE metabolism and disposition in mouse skin are different from those of BaP and indicate that the relatively low tumorigenicity of BaP-7,8-diol and anti-BPDE in mouse skin may be partially attributable to differences between the disposition of these metabolites when topically applied compared to when they are generated intracellularly from BaP.