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Stopped flow kinetics of pyrene transfer between human high density lipoproteins.

The transfer of pyrene between high density lipoproteins was studied as a model of lipid exchange. When high density lipoprotein containing pyrene was mixed with unlabeled lipoprotein, pyrene excimer fluorescence decreased with a half-time of approximately 3 ms. The rate of pyrene transfer was invariant over a 100-fold range of unlabeled lipoprotein concentrations. Since a decrease in excimer fluorescence indicates a decrease in the microscopic concentrations of pyrene, the observed fluorescence change relfects pyrene transfer to unlabeled lipoproteins, and, therefore, dilution of the pyrene molecules. When high density lipoprotein labeled with pyrene was rapidly diluted 1:14 into buffer, a small decrease in excimer fluorescence was observed. The half-time of this fluorescence change was also about 3 ms and represents the half-time for the dissociation of pyrene from high density lipoprotein into water. The latter observation, coupled with the invariant exchange rate with lipoprotein concentration suggests strongly that the limiting step in the transfer of pyrene between high density lipoproteins is the dissociation of pyrene into solvent. Finally, regardless of mechanism, the exchange of pyrene, and presumably other hydrophobic aromatic compounds, among serum high density lipoproteins is extremely fast. This result indicates that these types of compounds can be rapidly assimilated and transported through the body by plasma lipoproteins.

Binding Sites↗

Effects of butylated hydroxyanisole on the metabolism of benzo(a)pyrene by mouse lung microsomes.

Butylated hydroxyanisole (BHA) is a commonly used food additive with demonstrated inhibitory action against chemical carcinogenesis in animals. In order to elucidate the mechanism of the anticarcinogenic action, the effects of BHA on benzo(a)pyrene (BP) metabolism were studied with lung microsomes from female mice. BHA treatment (0.5% in the diet for 7 days) inhibited BP metabolism and altered the ratios among different metabolites as analyzed by high-performance liquid chromatography. The treatment reduced the metabolic formation of 9,10-dihydroxy-9,10-dihydrobenzo(a)pyrene, but not the production of 3-hydroxybenzo(a)pyrene and trans-4,5-dihydroxy-4,5-dihydrobenzo(a)pyrene. Since the gross microsomal cytochrome P-450 content was not significantly affected by the treatment, the change of regioselectivity in BP metabolism was probably due to the alteration of cytochrome P-450 isozyme composition by dietary BHA. General and regioselective inhibition of BP metabolism was also observed when BHA was added to the lung microsomal incubation mixture. The formation of 9,10-dihydroxy-9,10-dihydrobenzo(a)pyrene and 9-hydroxybenzo-(a)pyrene was inhibited more severely than that of trans-4,5-dihydroxy-4,5-dihydrobenzo(a)pyrene and trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene, but the production of 3-hydroxy-benzo(a)pyrene was not inhibited. Dietary BHA treatment also decreased the microsomal metabolism of trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene to n-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene and r-7,t-8-dihydroxy-c-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene. Considering that the former diol-epoxide is a suspected ultimate carcinogen, the observed inhibitions of BP metabolism in the formation of diol-epoxides may be closely related to the anticarcinogenic action of BHA.

Animals↗

Fluorometric Studies of Pyrene Adsorption on Porous Crystalline Cellulose.

Pyrene crystals were physically mixed with either porous crystalline cellulose (PCC) or octa decyl siryl silica-80Tm (ODS). Solid-state fluorescence spectra of pyrene were analyzed to estimate the interaction between pyrene and porous materials. Pyrene monomer emission was observed at 398 nm immediately after being mixed with PCC, while pyrene crystals showed only excimer emission at 475 nm, indicating that the pyrene molecules adsorbed onto the PCC surface in a short period. For the PCC system containing 1.0% pyrene, long-term storage caused an increase in the intensity of excimer-like emission peak at 477 nm accompanied by a decrease in the intensity of monomer emission peak at 398 nm. For the ODS system containing 1.0% pyrene, spectrometric changes were similar to those for the PCC system. In the process of interaction formation between pyrene and an additive, a two-step mechanism was proposed, i.e., the adsorption of pyrene molecules onto the surface of porous additives, and the formation of a ground state dimer of pyrene. The formation of dimeric pyrene could be associated with the surface polarity of additives. Copyright 1998 Academic Press.

Journal Article↗

An enantiomeric interaction in the metabolism and tumorigenicity of (+)- and (-)-benzo[a]pyrene 7,8-oxide.

The (+)- and (-)-enantiomers of benzo[a]pyrene 7,8-oxide are hydrated stereospecifically at C-8 to (-)- and (+)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, respectively, by rat hepatic epoxide hydrolase. The (-)-enantiomer of benzo[a]pyrene 7,8-oxide is metabolized by microsomal epoxide hydrolase at a rate 3- to 4-fold greater than the (+)-enantiomer. At low conversion of racemic substrate, however, benzo[a]pyrene 7,8-oxide is metabolized to the dihydrodiol at a rate equal to that of the (+)-enantiomer. An analysis of the enantiomeric composition of the dihydrodiol formed from the racemic substrate revealed preferential formation of (-)-trans-7,8-dihydroxy-7,8-dihydrobenzo-[a]pyrene. At low substrate conversion (< 20% metabolism), the enantiomeric purity of the dihydrodiol was much higher than at high substrate conversion (> 50% metabolism). Similar results were obtained with microsomes from hamster, rabbit, guinea pig, mouse, and human liver. These results indicate that epoxide hydrolase has a higher affinity for (+)-benzo[a]pyrene 7,8-oxide than for the (-)-enantiomer. The kinetics of hydration of (+)- and (-)-benzo[a]pyrene 7,8-oxide by purified epoxide hydrolase in detergent solution showed the (+)- and (-)-enantiomers to have apparent Km values of 1.7 and greater than or equal to 20 microM, respectively. Tumorigenicity studies with benzo[a]pyrene 7,8-oxide on mouse skin and in newborn mice revealed that (+)-benzo[a]pyrene 7,8-oxide, the metabolic precursor of the more tumorigenic (-)-7,8-dihydrodiol, is significantly more tumorigenic than the (-)-enantiomer. However, racemic benzo[a]pyrene 7,8-oxide was more tumorigenic than either enantiomer alone, indicating an enantiomeric synergism in the carcinogenicity of benzo[a]pyrene 7,8-oxide. The data are discussed in relation to the complete sequence of metabolic pathways leading to an ultimate carcinogen from benzo[a]pyrene.

Animals↗

Tumorigenicity of the diastereomeric bay-region benzo(e)pyrene 9,10-diol-11,12-epoxides in newborn mice.

The tumorigenic activities of benzo(e)pyrene, 9,10-dihydrobenzo(e)pyrene, 9,10-epoxy-9,10,11,12-tetrahydrobenzo(e)-pyrene, the diastereomeric bay-region 9,10-dihydroxy-11,12-epoxy-9,10,11,12-tetrahydrobenzo(e)pyrenes, and the K-region benzo(e)pyrene 4,5-oxide were assessed in newborn mice, Swiss-Webster mice received a total dose of 0.7 mumol of compound divided into three i.p. injections of 0.1, 0.2, and 0.4 mumol on the first, eighth, and 15th days of life, respectively. 9,10-Epoxy-9,10,11,12-tetrahydrobenzo(e)pyrene was highly toxic to the newborn mice, and the first injection of 0.1 mumol of this benzo(e)pyrene derivative killed all the mice within two weeks. The total dose of 9,10-epoxy-9,10,11,12-tetrahydrobenzo(e)pyrene was therefore reduced to 0.07 mumol in divided doses of 0.01, 0.02, and 0.04 mumol. When the animals were killed at 39 to 43 weeks of age, one of the diastereomeric bay-region diol-epoxides, (+/-)-9 beta, 10 alpha-dihydroxy-11 beta, 12 beta-epoxy-9,10,11,12-tetrahydrobenzo(e)pyrene, produced a small but significant increase in pulmonary tumors in male mice but had no significant hepatotumorigenic activity. The diastereomerically related diol-epoxide. (+/-)-9 beta, 10 alpha-dihydroxy-11 alpha, 12 alpha-epoxy-9,10,11,12-tetrahydrobenzo(e)pyrene, produced a significant incidence of hepatic tumors but had no effect on the formation of pulmonary tumors. Benzo(e)pyrene and the other benzo(e)pyrene derivatives were all nontumorigenic at the doses tested.

Animals↗

Removal of pyrene from contaminated sediments by mangrove microcosms.

The potential of mangrove wetland systems to remove pyrene from surface- or bottom-contaminated sediments was investigated by microcosm studies. The performance of two mangrove plant species, Kandelia candel and Bruguiera gymnorrhiza in pyrene removal was also compared. During the six-months experimental period, the growth of both species in the surface-contaminated microcosms was not significantly different from that in the bottom-contaminated ones, and was comparable to the control (without any pyrene contamination). At the end of six-months treatment, pyrene concentrations in contaminated sediments declined from an initial 3 microg g(-1) to less than 0.4 microg g(-1), indicating that pyrene was successfully removed by mangrove microcosms. Around 96.4% and 92.8% pyrene in microcosms planted with K. candel were removed from the surface- and bottom-contaminated sediments, respectively. The removal percentages were slightly lower in microcosms planted with B. gymnorrhiza. Significant accumulation of pyrene in roots was only found in microcosms having bottom-contaminated sediments, and pyrene concentrations were 3.05 microg g(-1) and 4.50 microg g(-1) in roots of K. candel and B. gymnorrhiza, respectively. These values were much higher than that in control microcosms (without pyrene contamination, root pyrene concentrations were 0.27 microg g(-1) for K. candel and 0.34 microg g(-1) for B. gymnorrhiza) and in microcosms with contaminated sediments placed at the surface layer. Nevertheless, the overall contribution of root accumulation and plant uptake to the removal of pyrene from contaminated sediments was insignificant.

Biomass↗

Cold and drought stress in combination with pyrene exposure: studies with Protaphorura armata (Collembola: Onychiuridae).

Adult survival of the springtail Protaphorura armata exposed to pyrene, a common soil pollutant, was investigated in combination with cold and drought stress, in three separate experiments. (1) A drought stress imposed subsequent to pyrene exposure in soil resulted in a significant decrease in springtail survival, when compared with controls exposed to pyrene and subsequently to 100% relative humidity. (2) A previous exposure to drought stress resulted in slightly improved survival of pyrene exposure at a concentration of 10mg/kg, but not at higher pyrene concentrations. When comparing tests 1 and 2, better survival was found in the latter test. When comparing the drought survival of springtails that had been previously exposed to pyrene with drought survival of springtails with no previous history of pyrene exposure, survival was significantly lower in the former. (3) Springtail survival of pyrene exposure was investigated at several temperatures. Springtails showed a significant improvement in survival at temperatures fluctuating between +1 degrees C and -1 degrees C in 12:12-h cycles, and at a constant -3 degrees C, at the highest pyrene concentration (300 mg/kg), while survival remained the same at all temperatures when springtails were exposed to lower pyrene concentrations. It is concluded that temperature and water availability are important factors when assessing the springtails' susceptibility to pyrene exposure.

Animals↗

Effect of environmental tobacco smoke on the metabolism of (-)-trans-benzo[a]pyrene-7,8-dihydrodiol in juvenile ferret lung and liver.

To evaluate the effects of "environmental tobacco smoke" (ETS) on developing lungs, juvenile ferrets were exposed to ETS at an average total particulate concentration of 381 +/- 97 mg/m3 for 2 h at the breathing zone. Twenty-four hours after the exposure, the ferrets were sacrificed and the metabolism of (-)-trans-benzo[a]pyrene-7,8-dihydrodiol was studied in the lung and liver homogenates. The rate of conversion of (-)-trans-benzo[a]pyrene-7,8-dihydrodiol to the ultimate carcinogen (+)-anti-benzo[a]pyrene-7,8-dihydrodiol-9,10- epoxide was twofold higher in the liver than that observed in the lung of control ferrets. After ETS exposure, the formation of free benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide was increased by 62% in the lung (p < .01). The DNA-bound metabolites were significantly increased only in the lung, while protein-bound metabolites were significantly increased in the liver after ETS exposure. Although glutathione conjugates tended to be increased both in the lung and liver, sulfate conjugates were significantly decreased in the lung after ETS exposure (p < .05). (+)-trans-Benzo[a]pyrene-7,8-dihydrodiol was used to study the relative contributions of cytochrome P-450 and peroxyl radical-mediated formation of benzo[a]-pyrene-7,8-dihydrodiol-9,10-epoxide. Peroxyl radical- and P-450-mediated conversion of (+)-trans-benzo[a]pyrene-7,8-dihydrodiol to benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide was proportionately equal in the ferret lung, whereas in the liver the P-450-mediated pathway was predominant. After ETS exposure there was a tendency for P-450-mediated formation of benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide to increase. These results demonstrate significant differences in the metabolism of (-)-trans-benzo[a]pyrene-7,8-dihydrodiol by the lung and liver of juvenile ferrets and suggest a significant role of peroxyl radical-mediated formation of (+)-anti-benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide in the lung, which may help explain discrepancy between the levels of P-450 and amounts of DNA adducts of polycyclic aromatic hydrocarbons in different organs in smokers.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

Rapid mineralization of benzo[a]pyrene by a microbial consortium growing on diesel fuel.

A microbial consortium which rapidly mineralized the environmentally persistent pollutant benzo[a]pyrene was recovered from soil. The consortium cometabolically converted [7-(14)C]benzo[a]pyrene to (14)CO(2) when it was grown on diesel fuel, and the extent of benzo[a]pyrene mineralization was dependent on both diesel fuel and benzo[a]pyrene concentrations. Addition of diesel fuel at concentrations ranging from 0.007 to 0.2% (wt/vol) stimulated the mineralization of 10 mg of benzo[a]pyrene per liter 33 to 65% during a 2-week incubation period. When the benzo[a]pyrene concentration was 10 to 100 mg liter(-1) and the diesel fuel concentration was 0.1% (wt/vol), an inoculum containing 1 mg of cell protein per liter (small inoculum) resulted in mineralization of up to 17.2 mg of benzo[a]pyrene per liter in 16 days. This corresponded to 35% of the added radiolabel when the concentration of benzo[a]pyrene was 50 mg liter(-1). A radiocarbon mass balance analysis recovered 25% of the added benzo[a]pyrene solubilized in the culture suspension prior to mineralization. Populations growing on diesel fuel most likely promoted emulsification of benzo[a]pyrene through the production of surface-active compounds. The consortium was also analyzed by PCR-denaturing gradient gel electrophoresis of 16S rRNA gene fragments, and 12 dominant bands, representing different sequence types, were detected during a 19-day incubation period. The onset of benzo[a]pyrene mineralization was compared to changes in the consortium community structure and was found to correlate with the emergence of at least four sequence types. DNA from 10 sequence types were successfully purified and sequenced, and that data revealed that eight of the consortium members were related to the class Proteobacteria but that the consortium also included members which were related to the genera Mycobacterium and Sphingobacterium.

Benzo(a)pyrene↗

Comparison of benzo(a)pyrene metabolism in bronchus, esophagus, colon, and duodenum from the same individual.

The metabolism of benzo(a)pyrene has been investigated in cultured normal human bronchus, colon, duodenum, and esophagus obtained from the same patient. The highest total metabolism was found in bronchus and duodenum, while the highest mean binding level was observed in the bronchus followed, in order, by the esophagus, duodenum, and transverse colon. A 30-fold interindividual variation in the binding level was found in each of the four organs studied, and a positive correlation between the binding levels in bronchus, colon, and duodenum was found. In human bronchus, a positive correlation was found between level of binding of benzo(a)pyrene to DNA and the amount of both benzo(a)pyrene 7,8-diol and the combined group of 3-hydroxybenzo(a)pyrene, benzo(a)pyrene 9,10-diol, and water-soluble metabolites. A significantly higher relative amount of benzo(a)pyrene tetrols and benzo(a)pyrene 9,10-diol was formed by human bronchus compared to the gastrointestinal tissues, while a higher level of benzo(a)pyrene phenols was formed by the latter. The relative distribution of benzo(a)pyrene-DNA adducts was similar in all four organs, the major DNA adduct being formed by trans-addition of anti-7,8-dihydroxy-9,10-epoxide-7,8,9,10-tetrahydrobenzo(a)pyrene to the 2-amino group at guanine. These results indicate that the metabolism of benzo(a)pyrene by at least four different organs is qualitatively similar but that quantitative differences exist.

Adolescent↗

Immunochemical study on the contributions of two molecular species of microsomal cytochrome P-450 to the metabolism of benzo(a)pyrene by rat liver microsomes.

The roles of two species of cytochrome P-450, the major cytochrome P-450 components of liver microsomes of phenobarbital-treated rats (PB-P-450) and 3-methylcholanthrene-treated rats (MC-P-448), were studied in the metabolism of benzo(a)pyrene in rat liver microsomes in vitro. Benzo(a)pyrene was incubated with polychlorinated biphenyl-treated rat liver microsomes, in which PB-P-450 and MC-P-448 constituted about 45 and 24% of the total cytochrome P-450, respectively. Then the metabolites were separated into those soluble in ethyl acetate and in water, and those covalently bound to protein. Using high-pressure liquid chromatography, the ethyl acetate-soluble metabolites were separated into three major groups, phenols, quinones, and dihydrodiols, including peaks of three unknown materials. Addition of anti-MC-P-448 immunoglobulin to the reaction mixture completely inhibited the formation of all ethyl acetate-soluble metabolites. In contrast, anti-PB-P-450 immunoglobulin did not inhibit the formations of 4,5-dihydro-4,5-dihydroxybenzo(a)pyrene and 3-hydroxybenzo(a)pyrene; partially inhibited the formations of 7,8-dihydro-7,8-dihydroxybenzo(a)pyrene, 9, 10-dihydro-9, 10-dihydroxybenzo(a)pyrene, and the three unknown materials; and caused 30 to 40% enhancement of the formations of 9-hydroxy-benzo(a)pyrene and benzo(a)pyrene-3,6-dione and 80% enhancement of that of benzo(a)pyrene-1,6-dione. Antibody against MC-P-448, but not against PB-P-450, also caused 75% inhibition of the formation of water-soluble metabolites and 85% inhibition of formation of benzo(a)pyrene metabolites covalently bound to protein. These results show that MC-P-448 is important in the metabolism of benzo(a)pyrene.

Animals↗

Role of Kupffer cells in storage and metabolism of benzo(a)pyrene in the liver.

This study investigates the possible role of Kupffer cells in storage and metabolism of benzo(a)pyrene in the liver. In perfused liver, benzo(a)pyrene (4-120 microM) in 0.3% albumin increased fluorescence (366-->405 mm) on the liver surface in a dose-dependent manner, suggesting that it accumulated in liver tissue. The maximal increase of benzo(a)pyrene fluorescence was diminished by 60% when Kupffer cells were destroyed by gadolinium chloride treatment (10 mg/kg iv). Gadolinium chloride also decreased the yield of isolated nonparenchymal cells by 65%. In frozen sections of livers perfused with 4 microM benzo(a)pyrene for 1 hr, fluorescence was approximately 5 times greater in cells lining the sinusoids than in parenchymal cells. Moreover, yellow-green fluorescent particles were detected in cultured Kupffer cells, but were barely visible in parenchymal and Ito cells, indicating that Kupffer cells actively accumulated benzo(a)pyrene. In contrast to the cell specificity for benzo(a)pyrene accumulation, rates of monooxygenation of benzo(a)pyrene were up to 20-fold higher in isolated parenchymal than in Kupffer cells. In nonparenchymal cells, basal rates of production of benzo(a)pyrene phenols were approximately 50 pmol/10(6) cells/hr. In contrast, rates were approximately 335 pmol/10(6) cells/hr in parenchymal cells. Further, total [3H]benzo(a)pyrene metabolism was approximately 8-fold higher in parenchymal than in nonparenchymal cells. Albumin increased production of benzo(a)pyrene phenols by 3-fold in parenchymal cells, but was without effect in nonparenchymal cells. Pretreatment of rats with gadolinium chloride increased the production of benzo(a)pyrene phenols in perfused liver by > 50%.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Neonatal modulation of adult rat hepatic microsomal benzo[a]pyrene hydroxylase activities by Aroclor 1254 or phenobarbital.

The constitutive and Aroclor 1254-induced activities of hepatic microsomal benzo[a]pyrene hydroxylases in male and female rats were determined in animals from ages 11 to 120 days. In 11-day-old noninduced male rats, benzo[a]pyrenediones and 9-hydroxybenzo[a]pyrene were the major microsomal metabolites; in 21-day-old males benzo[a]pyrene-diones and benzo[a]pyrene-9,10-dihydrodiol were predominant. In 60- and 120-day-old animals 3-hydroxybenzo[a]pyrene was the major microsomal metabolite. A similar trend was observed for the development of benzo[a]pyrene hydroxylase activities in female rats. With the exception of 4,5-dihydrodiol formation, the highest induction of individual and total benzo[a]pyrene hydroxylase activities by Aroclor 1254 was observed in the 21-day-old immature male rats, in which there was a 330- and 4.5-fold increase in the formation of 3-hydroxybenzo[a]pyrene and quinone metabolites, respectively. The induction of benzo[a]pyrene total metabolite formation by Aroclor 1254 in female rats from 11 to 120 days of age was relatively constant (i.e., 13.3- to 10.1-fold induction); however, the relative induction of the individual benzo[a]pyrene hydroxylases was highly variable. In a second set of experiments, male and female rats were neonatally exposed to phenobarbital (600 mumol/kg) or Aroclor 1254 (100 mumol/kg), and the effects of these xenobiotics on neonatal imprinting of hepatic microsomal benzo[a]pyrene hydroxylase activities were determined in the 120-day-old animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

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.

Administration, Cutaneous↗

(+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo (a)pyrene: a potent skin carcinogen when applied topically to mice.

(+/-)-trans-7,8-Dihydroxy-7,8-dihydrobenzo[a]-pyrene, a known metabolite of benzo [a]pyrene, has been tested for carcinogenic activity on mouse skin by topical application of 0.15 or 0.30 mumol every 2 weeks for 60 weeks. At the low dose (0.15 mumol), the compound was equipotent to the parent hydrocarbon, benzo[a]pyrene, and considerably more potent than its metabolic precursor, benzo[a]pyrene 7,8-oxide, in eliciting tumors, as determined by both the onset of tumors and the total number of animals developing carcinomas. Application of 7,8-epoxy-,8,9,10-tetrahydrobenzo[a]pyrene (0.30 mumol every 2 weeks), a compound related to the carcinogenic benzo[a]pyrene 7,8-oxide but with the double bond removed from the 9,10-position of the molecule, did not elicit any tumors. The above results indicate that the (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene is a more proximate carcinogen than benzo[a]pyrene 7,8-oxide and that the carcinogenicity of benzo[a]pyrene 7,8-oxide and (+/-)trans-7,8-digydrobenzo[a]pyrene may be due to metabolic conversion of these compounds to the highly reactive and mutagenic stereoisomers of 7,8-digydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene.

Animals↗

High-pressure liquid chromatography analysis of benzo(a)pyrene metabolism by microsomal enzymes from rhesus liver and lung.

The metabolism of benzo(a)pyrene was determined, using rhesus monkey hepatic and pulmonary microsomal enzymes. Metabolites were separated by high-pressure liquid chromatography and identified using known reference standards. Metabolites were quantitated by scintillation spectrometry. Both liver and lung microsomes metabolized benzo(a)pyrene to the following metabolites: 9,10-, 7,8-, and 4,5-dihydrodihydroxybenzo(a)pyrene; benzo(a)pyrene-1,6-dione, -3,6-dione, and -6,12-dione; and 9- and 3-hydroxybenzo(a)pyrene. Two unidentified metabolites and one metabolite region which chromatographed prior to 9,10-dihydrodihydroxybenzo(a)pyrene were produced by both liver and lung microsomes. The two unknown peaks were located between, 9,10- and 4,5-dihydrohidroxybenzo(a)pyrene. Two additional unknown metabolites were produced only in the liver and had retention times slightly greater than the 4,5- and 7,8-dihydrodihydroxybenzo(a)pyrene metabolites, respectively. Quantitative determination of benzo(a)pyrene metabolism revealed large differences for the three monkeys and the respective tissue activities. Liver activity for each animal was substantially higher than lung activity for all benzo(a)pyrene metabolites. The ratio of the metabolites also differed between the liver and lung. 3-Hdyroxybenzo(a) pyrene represented over 60% of the total liver metabolite fraction and 30% of the total lung metabolite fraction. The total quinone fraction represented between 7 and 13% of the total metabolites in the liver and comprised over 40% of the total lung metabolites. The metabolite ratios for the dihydrodiols were very similar for both tissues.

Animals↗

Oxidation of benzo[a]pyrene by the filamentous fungus Cunninghamella elegans.

Cunninghamella elegans oxidized benzo[a]pyrene to several metabolic products. Compounds that were isolated and identified were: trans-9,10-dihydroxy-9,10-dihydrobenzo[a]pyrene, trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, benzo[a]pyrene 1,6-quinone, benzo[a]pyrene 3,6-quinone, 9-hydroxybenz[a]pyrene, and 3-hydroxybenzo[a]pyrene. In addition, an unidentified dihydroxybenzo[a]pyrene metabolite was also formed. Experiments with [14C]benzo[a]pyrene showed that over a 96-h period, 18.4% of the hydrocarbon was converted to metabolic products. Most of the metabolites were sulfate conjugates as demonstrated by the formation of benzo[a]pyrene quinones and phenols after treatment with aryl sulfatase. Glucuronide and sulfate conjugates were also detected as water-soluble metabolites. The results show that benzo[a]pyrene is metabolized by a filamentous fungus in a manner that is remarkably similar to that observed in higher organisms.

Benzopyrenes↗

Tumorigenic activity of benzo(e)pyrene derivatives on mouse skin and in newborn mice.

The tumorigenic activities of benzo(e)pyrene and several of its derivatives were determined in two mouse tumor models. Newborn Swiss-Webster mice were given i.p. injections of 0.4, 0.8, and 1.6 mumol of compound on the first, eighth, and 15th day of life, respectively. When the mice were 62 to 66 weeks old, the experiment was terminated by killing the animals. Benzo(e)pyrene, trans-4,5-dihydroxy-4,5-dihydrobenzo(e)pyrene, and trans-9,10-dihydroxy-9,10-dihydrobenzo(e)pyrene had little or no tumorigenic activity in lung tissue, although trans-9,10-dihydroxy-9,10-dihydrobenzo(e) pyrene did induce a significant number of hepatic tumors. The tumor-initiating activities of benzo(e)pyrene and several of its derivatives were determined on the skin of female CD-1 mice. A single topical application of 1.0 to 6.0 mumol of the test compound was followed 7 days later by twice-weekly applications of the tumor promoter 12-O-tetradecanoylphorbol-13-acetate for 35 weeks. Control mice and mice treated with 6.0 mumol of benzo(e)pyrene, trans-4,5-dihydroxy-4,5-dihydrobenzo(e)pyrene, trans 9,10-dihydroxy-9,10-dihydrobenzo(e)pyrene, and trans-9,10-dihydroxy-9,10,11,12-tetrahydrobenzo(e)pyrene had a tumor incidence of less than 20% and had less than or equal to 0.25 papillomas/mouse. 9,10-Dihydrobenzo(e)pyrene was the only derivative tested that had significant tumor-initiating activity on mouse skin; an initiating dose of 2.5 mumol gave a 67% tumor incidence and 1.43 papillomas/mouse.

Animals↗