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PubMed1983

Protective effects of ellagic acid and other plant phenols on benzo[a]pyrene-induced neoplasia in mice.

The inhibitory effects of three phenolic compounds (ferulic, chlorogenic and ellagic acids) on benzo[a]pyrene- and 7,12-dimethylbenz[a]anthracene-induced neoplasia have been investigated in mice. Ellagic acid was the most potent antagonist of tumorigenesis since this compound is active, by i.p. administration or added in the diet, on benzo[a]pyrene-induced pulmonary adenoma formation in A/J mice and, after topical application, on 7,12-dimethylbenz[a]anthracene-induced skin tumorigenesis in NMRI Swiss mice. If ellagic acid has little or no effect on the number of tumor bearing animals, the incidence of pulmonary tumors per animal is decreased by greater than 50%. Ferulic acid and chlorogenic acid (5 X 100 mg/kg, by i.p. route) were also active, but less than ellagic acid, against the lung carcinogenesis by benzo[a]-pyrene (100 mg/kg, i.p.) but were totally ineffective against the formation of skin tumors by 7,12-dimethylbenz[a]anthracene. These results remarkably paralleled the in vitro antimutagenic effects of these compounds shown by Wood et al. on benzo[a]pyrene. It must be noted that ellagic acid only exerted, by i.p. route, a severe toxicity after four injections of 100 mg/kg, in oil suspension, whereas the oral administration in the diet (a daily dose of 100 mg/kg during 15 days) did not cause any toxicity.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1983

Carcinogenic activity of benzo[a]pyrene and some of its synthetic derivatives by direct injection into the mouse fetus.

We have studied the carcinogenic effects of direct injection of benzo[a]pyrene (BP) and 6-methylbenzo[a]pyrene (BP6M) into fetal Swiss mice at 15 days of intrauterine growth. To determine the sensitivity of mouse fetuses to the action of ultimate carcinogens, benzo[a]pyrene-4,5-oxide (BPO) and a racemic mixture of 7 beta,8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9, 10-tetrahydrobenzo[a]pyrene (BPDE), respectively, a proximal metabolite and an ultimate carcinogenic metabolite of BP, were also investigated in our system. The compounds were dissolved in acetone and trioctanoin (1:1) and injected at doses ranging from a minimum of 0.4 to a maximum of 19.8 nmol/fetus. Controls received 1 microliter/fetus of solvent. The experiment was terminated when the animals were 12-15 weeks old. On the basis of the percentages of lung adenomas and the average number of nodules/mouse, BPDE appears to be the most potent carcinogen of this group, causing 55% of tumors at a dose of 0.4 nmol and 73% at a dose of 4.0 nmol/fetus. At the dose of 0.4 nmol/fetus, neither BP nor BP6M induced tumors; injection of 4.0 or 19.8 nmol/fetus, caused incidences of 53% and 92%, respectively, in the case of BP6M and 24% and 39%, respectively, in the case of BP. BPO was not tumorigenic in this system, even at doses as high as 15.7 nmol/fetus. Eleven percent of the control animals had lung adenomas. The average number of nodules/mouse varied with the compound and the dose injected and closely followed the pattern of tumor incidence.

Animals↗

PubMed1985

An applied synchronous fluorescence spectrophotometric assay to study benzo[a]pyrene-diolepoxide-DNA adducts.

Synchronous scanning fluorescence with a fixed wavelength difference (delta lambda) of 34 nm between excitation and emission was used to quantitate benzo[a]pyrene-diol epoxide (BPDE)-DNA adducts. Fluorescence emission maxima occurred at 382 nm for BPDE-DNA and at 379 nm for benzo[a]pyrene-tetrols and -triol, which are hydrolysis products of BPDE. Similarly, the peak for pyrene was at 372 nm and for 1-nitropyrene at 386 nm. The minimum detectable amount of BPDE-moieties in in vitro modified BPDE-DNA, after hydrolysis in HCl, was 20 fmol in 100 micrograms of DNA, which is equivalent to 1 adduct per 1.4 X 10(7) nucleotides. The correlation of fluorescence intensity and the amount of BPDE-moieties was linear between 20 fmol and 1 pmol. DNA isolated from human lymphoblasts incubated with BPDE had the same fluorescence peak and the correlation between the fluorescence intensity and the amount of BPDE in the incubation mixture was linear. Among the DNA-samples from peripheral blood lymphocytes of 30 aluminum plant workers, only one sample was found to contain a peak similar to BPDE-DNA. None of the DNA-samples from 10 persons not occupationally exposed were positive. Measurement of BPDE-DNA adducts by synchronous fluorescence spectrophotometry should be useful in monitoring human exposure to benzo[a]pyrene.

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

PubMed1986

The blocking effect of disodium cromoglycate on carcinogenesis induced by benzo[a]pyrene.

Subcutaneous injection of 3 mg benzo[a]pyrene into the right hind leg of experimental rats resulted in local tumors in the area of application in all the animals. The same dose of disodium cromoglycate applied prior to benzo[a]pyrene prevented tumor formation or provoked a significant inhibition of the carcinogenic process, e.g. delay of tumor formation and of a reduced size, a number of mitoses and cytological abnormalities and a prolongation of the animal's life. In the in vitro Salmonella typhimurium mutation assay of Ames, disodium cromoglycate eliminated toxicity and mutagenicity of 5-nitro-2-furylacrylic acid. The results would suggest that the initial response of the cell to benzo[a]pyrene had been conducted through the calcium-dependent exocytic pathway, which can be efficiently blocked by disodium cromoglycate, probably by preventing the signals increase in free intracellular calcium concentration. The preserved calcium balance as a consequence of the effect of disodium cromoglycate on benzo[a]pyrene-induced cell response might be a factor responsible for the carcinogenesis inhibition phenomena. The results of the in vitro study in correlation with those obtained in vivo support the suggestion that disodium cromoglycate may influence parallel structures in various cell types.

Acrylates↗

PubMed1987

Sulfite-dependent mutagenicity of benzo[a]pyrene derivatives.

Benzo[a]pyrene (BP) and sulfur dioxide (SO2) are ubiquitous air pollutants and are also components of tobacco smoke. Although SO2 itself is not carcinogenic, concurrent administration with BP results in enhancement of respiratory tract tumorigenesis. In biological systems, SO2 exists as its hydrated form, sulfite (SO3(2-) ). Sulfite readily undergoes autoxidation, generating potent oxidant species. When 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol) is included in sulfite autoxidation mixtures it is converted to more polar products, most notably 7,8,9,10-tetrahydroxy-7,8,9,10-tetrahydrobenzo[a]pyrenes (BP tetraols). This implies the intermediacy of 7,8-dihydroxy-9,10-epoxy- 7,8,9,10-tetrahydro-benzo[a]pyrenes (BPDE). We report here the sulfite-dependent conversion of BP-7,8-diol to forms highly mutagenic to Salmonella typhimurium strain TA 98. This activation is observed at BP-7,8-diol concentrations of from 2 to 40 microM and at sulfite concentrations of from 0.5 to 10 mM. In the presence of 10 microM BP-7,8-diol, half-maximal activation is observed at 1.6 mM sulfite. Sulfite itself is neither toxic nor mutagenic to the bacteria under these conditions. The time course of the activation of BP-7,8-diol and its sensitivity to inhibition by antioxidants indicate a requirement for sulfite autoxidation. These data further support the sulfite-dependent epoxidation of BP-7,8-diol. Not only does sulfite convert this promutagen to its active mutagenic form, sulfite also enhances the mutagenic activity of BP diolepoxides toward the tester strain. The reversion frequency in response to 0.1-0.5 microM anti-BPDE is increased by up to 33% in the presence of 1 mM sulfite, and by up to 270% with 10 mM sulfite. The mechanism of this enhancement of anti-BPDE activity is not known, but could be related to inhibition of the glutathione-S-transferase system which has been previously reported for sulfite. These results are discussed in regard to the noted cocarcinogenicity of sulfur dioxide for BP.

Benzo(a)pyrene↗

PubMed1995

Induction of micronuclei in metabolically competent rat hepatoma cell lines by the promutagens 7,12-dimethylbenz[a]anthracene, benzo[a]pyrene and cyclophosphamide.

The capability of two rat hepatoma cell lines, H4IIEC3/G- and 2sFou, to detect genotoxicity of xenobiotics, was evaluated in a micronucleus assay. In this system, the cells act as the activation source as well as the target cells for the DNA damage. The study was performed using 7,12-dimethylbenz[a]anthracene, benzo[a]pyrene and cyclophosphamide, as pro-mutagens and mitomycin C as a direct acting mutagen. In both cell lines a significant micronucleus induction was observed after exposure to each test compound, starting from 25 nM for 7,12-dimethylbenz[a]anthracene, 8 microM for benzo[a]pyrene, 0.5 mM for cyclophosphamide and 0.4 microM for mitomycin C. A period of 24 h treatment and 48 h growth was sufficient for induction and expression of micronuclei. The two hepatoma lines behave in a similar way with regard to the pro-mutagen activation. The results obtained in this study with these differentiated hepatoma lines demonstrate that they are metabolically competent to activate the promutagens 7,12-dimethylbenz[a]anthracene, benzo[a]pyrene and cyclophosphamide into their biologically active metabolites as measured by micronucleus induction in our experiments. However, the variable dose responses to 7,12-dimethylbenz[a]anthracene and benzo[a]pyrene in some of the repeated experiments, suggest unstable activity of enzymes involved in polycyclic aromatic hydrocarbons metabolism in these cell lines.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2001

Micronucleus induction and chromosomal aberration of 1- and 3-nitroazabenzo[a]pyrene and their N-oxides.

Nitro-azabenzo[a]pyrenes, 1- or 3-nitro-azabenzo[a]pyrene and their N-oxides are nitrated derivatives of azabenzo[a] pyrene (ABP) containing nitrogen in the 6-position of benzo[a]pyrene (B[a]P). The nitro-ABP-N-oxides (ABPOs) were formed by reaction of ABP with excess HNO(3). These derivatives were noteworthy as potent mutagens for Salmonella strains, and were present in fine particles of diesel particulates. In this study, micronucleus induction in mice and chromosomal aberrations due to means of Chinese hamster lung fibroblast (CHL) cells were investigated to determine genotoxicity in order to define the relationship with the mutagenic potency of these derivatives. The induction of micronucleus polychromatic erythrocytes (MNPCEs) was dependent on the dose response of 10-40 mg for 3-N-6-ABP, and of 10-40 mg for 1-N-6-ABP, and in addition, 1- and 3-N-6-ABPOs markedly induced MNPCEs in a dose range of 10-400 mg and from 1 to 80 mg, respectively, when the compound was intraperitoneally administrated in two mice at each dose. The results show that of the four compounds, 3-N-6-ABPO demonstrated a marked increase in MNPCES: On the other hand, chromosomal aberrations of the four compounds were investigated by the duplicate tests using CHLS: The results after a 48 h treatment induced aberrations of the chromatid type, chromatid breaks and exchanges for 1- and 3-N-6-ABP, and mainly chromatid exchanges for 1- and 3-N-6-ABPO. The frequency of chromosomal aberrations associated with nitro substitution on the ABPO structure. Chromosomal aberrations of nitro derivatives of ABPO substituted at the 3-position on the structure were more potent than those at the 1-postion. N-oxide derivatives have been found to be reduced to anion radicals much more easily than azaB[a]P and its nitro derivatives. This suggests that the electrochemical reduction of the chemicals plays an important role in the metabolic activation of nitrated B[a]P derivatives.

Animals↗

PubMed2004

Synthesis and properties of fluorescent-labeled oligonucleotides containing amide-linked dinucleoside with pyrene.

Oligonucleotides containing 2'-pyrene modified uridine (Upy) exhibit the increase of the pyrene monomer emission when hybridize with the complementary RNA. In order to examine the effect of restricted C3'-endoconformation of the Upy residue for the enhancement of the pyrene monomer emission, an oligodeoxyribonucleotide containing amide-linked dinucleoside with a pyrene group at the sugar residue (UapyODN) was synthesized by use of 3'-carboxymethyl-5'-O-dimethoxytrityl-2'-O-(1-pyrenylmethyl)-3'-deoxyuridine. NMR analysis indicated that the furanose ring of the uridine derivative was fixed to C3'-endo conformation. The UapyODN showed increase of fluorescence intensity to 7-fold at 375 nm when hybridized with complementary oligoribonucleotide. This result suggests that the UapyODN can detect the target RNA.

Amides↗

PubMed1994

X-ray diffraction study of the disorder-to-order transition at approximately 160 K in the pi-molecular compound pyrene ... pyromellitic dianhydride.

The crystal structure of pyrene ... pyromellitic dianhydride [(C16H10:C10H2O6); PYRPMA] has been studied over the range 300-19 K, using the low-temperature accessory designed by Samson, Goldish & Dick [J. Appl. Cryst. (1980), 13, 425-432] for a four-circle diffractometer. Earlier results for the disordered and ordered structures [Herbstein & Snyman (1969). Philos. Trans. R. Soc. London Ser. A, 264, 635-666] are confirmed and extended. At 295 K, a = 13.94 (1), b = 9.34 (1), c = 7.31 (1) A, beta = 93.65 (9) degrees, space group P2(1)/a, Z = 2, with pyrenes and pyromellitic dianhydrides (PMDAs) at crystallographic centres of symmetry. At 19 K, a = 13.664 (3), b = 9.281 (2), c = 14.420 (3) A, beta = 91.80 (2) degrees, space group P2(1)/n, Z = 4, with two sets of pyrenes at independent centres of symmetry and the four PMDAs at general positions. There are no discontinuities in cell dimensions with temperature (measurements at approximately 10 K intervals, down to 19 K) but db/dT and d beta/dT show discontinuities at approximately 167 K. Superlattice reflections appear below approximately 164 K (= Tc by X-ray diffraction), corresponding to the doubling of c and change of space group; the specific heat shows an anomalous increase over the range 120-155 K, giving a lambda-type peak. These results show that the transition is second order with regard to Ehrenfest's criteria. PYRPMA is a co-elastic crystal and quantitative analysis shows a linear dependence of the squares of spontaneous strain and of normalized superlattice intensity on temperature; hence, in terms of Landau theory, the transition is tricritical. However, the excess specific heat cannot be explained entirely on this basis. The physical nature of the transition is discussed. PYRPMA is so far unique among the pi-molecular compounds showing disorder-to-order transitions in the solid state in that there is a doubling of one of the axes; all indications are, however, that it resembles the other examples in the sense that subtle intermolecular packing interactions (here between pyrenes and PMDAs) are the driving force for the transition rather than electronic or charge-transfer interactions.

Benzoates↗

PubMed2000

Are the members of a paving crew uniformly exposed to bitumen fume, organic vapor, and benzo(a)pyrene?

The goal of this investigation was to assess if and when a crew of paving workers is uniformly exposed to bitumen fume, organic vapor, and benzo(a)pyrene. Data on paving workers with up to six repeated exposure measurements were extracted from a database of exposure measurements developed within a study of the European asphalt industry (N = 591). The uniformity of exposures to bitumen fume, organic vapor, and benzo(a)pyrene was evaluated while grouping individuals by job title, primary tasks, crew membership, and use of coal tar (discontinued in Western Europe). The estimated ranges within which 95% of individual mean exposures were expected to fall ((BW)R0.95) were used to assess exposure uniformity. Variance components were estimated by constructing mixed-effects models, with grouping variables as fixed effects and worker identity as random effect. The influence of duration of the sampling survey on estimates of exposure variability for a crew was also examined. There was a substantial variability in exposures between paving crews, as well as persons holding the same job or doing the same task, but each crew was uniformly exposed to bitumen fume and benzo(a)pyrene ((BW)R0.95 2 and 1, respectively). Workers within the same crew engaged in paving with coal tar-containing binders were not, however, uniformly exposed to benzo(a)pyrene. Also, organic vapor exposures were not uniform among the members of a paving crew ((BW)R0.95 = 15). Sampling campaigns of up to 7 months had little impact on the estimates of within- and between-worker variability. These findings should assist investigators studying paving operations in optimizing their sampling, exposure assessment, and risk evaluation protocols. The results support the notion that only empirically determined predictors of exposure can yield optimal grouping, unlike a priori grouping strategies based on general descriptors such as jobs title or tasks performed.

Air Pollutants, Occupational↗

PubMed1993

Aggregation of pyrene-labeled microsomal glutathione S-transferase. Effect of concentration.

Microsomal glutathione S-transferase was labeled by the fluorescence probe N-(1-pyrenyl)maleimide which modified 1 mol thiol residue/mol protein. The enzyme activity increased about tenfold after the binding. The pyrene-labeled microsomal glutathione S-transferase exhibited two fluorescence bands which are typical of pyrene; one at 393 nm attributable to unassociated pyrenes, the other at 480 nm attributable to pyrene excimers (excited dimers). The excimeric fluorescence increased at high protein concentrations indicating a shift of the equilibrium of labeled polypeptide chains from trimeric complexes, the functional unit of microsomal glutathione S-transferase, to larger aggregates. At 25 degrees C and at a 1% Triton X-100 concentration, the calculated equilibrium constant of this process is 65 microM. Along with the formation of large aggregates, a progressive increase of the enzymic activity was observed. Thus, N-(1-pyrenyl)maleimide appears to be a very useful probe to study the supramolecular structure of this enzyme.

Animals↗

PubMed2006

In-situ enumeration and probing of pyrene-degrading soil bacteria.

Inferences about which microorganisms degrade polycyclic aromatic hydrocarbons in contaminated soils have largely been obtained using culture-based techniques, despite the low percentage of microorganisms in soil that are believed to be culturable. We used a substrate-responsive direct viable count method to identify and quantify potential polycyclic aromatic hydrocarbon-degrading bacteria in a soil containing petroleum wastes. Bacteria were extracted and their response to substrates determined in the presence of DNA gyrase inhibitors, which cause viable and active cells to elongate. When yeast extract, a widely used carbon source, was added as a growth substrate, together with nalidixic acid, piromidic acid and ciprofloxacin, a significant increase in elongated cells to 47%, 37% and 22%, respectively, was observed within 24 h. With pyrene as the main substrate, 10 mg L(-1) of nalidixic acid or piromidic acid caused 18-22% and 8-12%, respectively, of the cells to elongate within 24 h; whereas the effect of 0.5 mg L(-1) ciprofloxacin was not significant until 53 h later. Enlarged cells were identified and enumerated by fluorescent in situ hybridization, using Alpha-, Beta- and Gammaproteobacteria, and domain Bacteria-specific probes. The Bacteria-specific probe detected 35-71% of the total microorganisms detected by the DNA-binding dye 4,6-diamidino-2-phenylindole. Initially, 44%, 13% and 5% of the total bacteria in the soil extract were Alpha-, Beta- and Gammaproteobacteria, respectively. Without pyrene or a gyrase inhibitor, these subgroups decreased to 30% of the total population but were predominant with piromidic acid or unchanged with ciprofloxacin when pyrene was the main substrate. The proportion of elongated Alpha- and Betaproteobacteria (potential pyrene degraders) increased significantly (P<0.05). This approach links phylogenetic information with physiological function in situ without the conventional cultivation of bacteria and can be used to probe and enumerate degradative groups at even a finer level of discrimination.

Anti-Bacterial Agents↗

PubMed1996

Evidence for cytochrome P-450 and P-450-mediated benzo(a)pyrene hydroxylation in the white rot fungus Phanerochaete chrysosporium.

The presence of cytochrome P-450 and P-450-mediated benzo(a)pyrene hydroxylase activity in both microsomal and soluble fractions of the white rot fungus Phanerochaete chrysosporium was shown. The reduced carbon monoxide difference spectrum showed maxima at 448-450 and 452-454 nm for microsomal and cytosolic fractions, respectively. Both P-450 fractions produced a Type I substrate binding spectrum on addition of benzo(a)pyrene. Activity for benzo(a)pyrene hydroxylation was NADPH dependent and inhibited by carbon monoxide. Km values for activity showed a difference between the cellular fractions with a Km of 89 microM for microsomal P-450 and 400 microM for cytosolic P-450. The Vmax values observed were 0.83 nmol min-1 (nmol microsomal P-450)-1 and 0.4 nmol min-1 (nmol cytosolic P-450)-1. The results indicate that P-450-mediated benzo(a)pyrene hydroxylase activity could play a role in xenobiotic transformation by this fungus beside the known ligninolytic exocellular enzymes.

Basidiomycota↗

PubMed1995

Cassia senna inhibits mutagenic activities of benzo[a]-pyrene, aflatoxin B1, shamma and methyl methanesulfonate.

Ethanol extract of Senokot tablets (Cassia senna concentrate used as vegetable laxative), was found to be non-mutagenic while it inhibited the mutagenicity of benzo[a]pyrene, shamma, aflatoxin B1 and methyl methanesulfonate in the Ames histidine reversion assay using the Salmonella typhimurium tester strain TA98. While the Senokot extract completely inhibited the mutagenicity of promutagens (i.e. metabolic activation dependent) like benzo[a]pyrene and shamma, it reduced the mutagenic activity of the direct acting mutagen methyl methanesulfonate by only 58%. The mutagen aflatoxin B1 showed a 25-fold increase in the number of histidine revertants per plate at low concentrations (1.0-4.0 micrograms/plate) in the presence of metabolic activation system while at high concentrations (10.0-30.0 micrograms/plate) it proved to be weakly mutagenic (with a 5-fold increase in the number of histidine revertants/plate) without metabolic activation. The Senokot extract completely inhibited the mutagenic effect of low concentrations of aflatoxin B1 in the presence of metabolic activation but not that resulting from higher concentrations without metabolic activation. The results obtained with benzo[a]pyrene, shamma and aflatoxin B1 indicated that the antimutagenic effects of Senokot extract could be largely due to an interaction with the metabolic process involved in the activation of procarcinogens. However, the results obtained with methyl methanesulfonate suggested that factors in Senokot may also interact with direct mutagens to produce some antimutagenic effects. An ethanol extract of crude senna leaves found to be weakly mutagenic also inhibited (though less than Senokot) the mutagenic effect of benzo[a]pyrene suggesting that the antimutagenic principle is present in the complex plant material itself.

Aflatoxin B1↗

PubMed1989

Fluorescence spectroscopy of benzo[a]pyrene diol epoxide-DNA adducts. Conformation-specific emission spectra.

The fluorescence characteristics of adducts derived from the covalent binding of the highly tumorigenic (+) and the non-tumorigenic (-) enantiomers of trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) to native calf thymus DNA are significantly different from one another both at room temperature and at 77 K. The ratio R of fluorescence intensities of the (0,0) band I (situated near 380 nm) and vibronic band V (near 400 nm) of the pyrene ring system in the BPDE-DNA adducts and of the tetraol (BPT) hydrolysis product of BPDE is very sensitive to the polarity of the solvent, thus mimicking the well known behavior of pyrene itself (A. Nakajima, 1971, Bull. Chem. Soc. Jpn. 44, 3272). The fluorescence excitation and emission spectra of the (+)-BPDE-DNA adducts are relatively sharp and only slightly red-shifted (2-3 nm) with respect to those of BPT in aqueous buffer solution, and R = 1.07 when the fluorescence is excited at the maximum of the absorption spectrum; this compares with R = 1.17 for BPT in water, R = 0.75 in ether, and R = 0.84 for noncovalently intercalated BPT. These results suggest that the pyrene ring system in the covalent (+)-BPDE-DNA adducts is located in an environment which is relatively exposed to the aqueous environment, while physically intercalated BPT molecules are located at hydrophobic binding sites. The fluorescence characteristics of the (-)-BPDE-DNA adducts are more heterogeneous and thus more complex than those of the (+)-adducts. The R ratio depends rather strongly on the wavelength of excitation; a minor, more highly fluorescent and relatively solvent-accessible form of adducts exhibits an R ratio of 1.01. The major, less solvent accessible form is characterized by a larger red shift in the absorption spectrum (approximately 10 nm) and emission spectrum (approximately 6 nm for the (0,0) band) relative to BPT, and an R ratio of 1.07. These characteristics suggest that the local environments of the pyrenyl residues in the (-)-BPDE-DNA adducts are significantly different from those of BPT bound noncovalently to DNA by the intercalation mechanism. Fluorescence methods, particularly at low temperatures where the bands are better resolved and the fluorescence yields are significantly greater than at room temperature, can also be used to distinguish covalent DNA adducts derived from the binding of (+)-BPDE and (-)-BPDE to native double-stranded DNA.

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

PubMed1989

"4S" polycyclic aromatic hydrocarbon binding protein. Its role as a benzo(a) pyrene cytosolic carrier to the microsomes of DBA/2N mouse liver.

Rodent liver cytosol and other biological systems contain two proteins that bind polycyclic aromatic hydrocarbons (PAH) in a non covalent manner and that sediment at a different rate when centrifuged on sucrose gradient. The role of the smaller protein ("4S" PAH-BP) was studied. When DBA/2N mouse liver homogenate was incubated with 3H-BaP, most of the radioactivity was found in the microsomal subcellular fraction. The cytosol binding activity apparently decreased but reincubation of the cytosol with the radioactive ligand completely restored "4S" PAH-BP activity. The microsomal uptake of 3H-BaP can be studied in a reconstituted system in which microsomes are incubated with radioactive benzo(a)pyrene in the presence of crude cytosol. In these conditions the microsomal uptake rate of 3H-BaP increased with the temperature and at 37 degrees C ten minutes were required to reach the plateau. When cytosol was substituted by HEDG buffer, the amount of radioactivity found in the microsomes decreased drastically. 0.2 microM was the benzo(a)pyrene concentration required to saturate the microsomes. When microsomes were incubated with ammonium sulfate cytosolic fractions or with homogeneously purified "4S" PAH-BP, the 3H-BaP uptake was restored and reached the maximum with 3 micrograms/ml of purified protein. The radioactive benzo(a)pyrene bound to microsomes was oxidated in the presence of NADPH regenerating system. The oxidated products were discharged from microsomes only when "4S" PAH-BP was either present during the incubation or added at its end. Thus, this protein is able to transfer benzo(a)pyrene to the microsomal metabolization sites and to facilitate the release of oxidized products and, presumably, bind them.

Ammonium Sulfate↗

PubMed1995

Evaluation of the generation of genotoxic and cytotoxic metabolites of benzo[a]pyrene, aflatoxin B1, naphthalene and tamoxifen using human liver microsomes and human lymphocytes.

1. The ability of model stable epoxides and metabolites generated by human liver microsomes from benzo[a]pyrene, aflatoxin B1, naphthalene and tamoxifen to produce cytotoxicity and genotoxicity in human peripheral lymphocytes has been investigated. 2. The stable epoxides 1,1,1 trichloropropene-2,3-oxide (100 microM) and trans stilbene oxide (100 microM) as well as metabolites generated from aflatoxin B1 (30 microM) and naphthalene (100 microM) by an extracellular metabolising system were toxic to isolated resting mononuclear leucocytes (MNLs), whereas glycidol (100 microM), benzo[a]pyrene (100 microM) and tamoxifen (50 microM) were not. 3. The stable epoxides 1,1,1 trichloropropene-2,3-oxide (100 microM) and trans stilbene oxide (100 microM) but not glycidol (100 microM) were toxic to dividing lymphocytes only after a 72-h exposure. Tamoxifen (30 microM), aflatoxin B1 (30 microM) and their metabolites were also toxic to dividing lymphocytes. Benzo[a]pyrene (100 microM) and naphthalene (100 microM) were not toxic either in the absence or presence of the extracellular metabolising system. 4. Benzo[a]pyrene (100 microM) and aflatoxin B1 (30 microM) were directly genotoxic to lymphocytes, this genotoxicity was significantly enhanced by the presence of the extracellular metabolising system. This indicates that both intracellular and extracellular bioactivation of these two compounds can produce genotoxicity. In contrast, naphthalene and tamoxifen were non-genotoxic.

1-Propanol↗

PubMed2006

Protonation of excited state pyrene-1-carboxylate by phosphate and organic acids in aqueous solution studied by fluorescence spectroscopy.

Pyrene-1-carboxylic acid has a pK of 4.0 in the ground state and 8.1 in the singlet electronic excited state. In the pH range of physiological interest (pH approximately 5-8), the ground state compound is largely ionized as pyrene-1-carboxylate, but protonation of the excited state molecule occurs when a proton donor reacts with the carboxylate during the excited state lifetime of the fluorophore. Both forms of the pyrene derivatives are fluorescent, and in this work the protonation reaction was measured by monitoring steady-state and time-resolved fluorescence. The rate of protonation of pyrene-COO(-) by acetic, chloroacetic, lactic, and cacodylic acids is a function of DeltapK, as predicted by Marcus theory. The rate of proton transfer from these acids saturates at high concentration, as expected for the existence of an encounter complex. Trihydrogen-phosphate is a much better proton donor than dihydrogen- and monohydrogen-phosphate, as can be seen by the pH dependence. The proton-donating ability of phosphate does not saturate at high concentrations, but increases with increasing phosphate concentration. We suggest that enhanced rate of proton transfer at high phosphate concentrations may be due to the dual proton donating and accepting nature of phosphate, in analogy to the Grotthuss mechanism for proton transfer in water. It is suggested that in molecular structures containing multiple phosphates, such as membrane surfaces and DNA, proton transfer rates will be enhanced by this mechanism.

Acids↗