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PubMed1994

Degradation of pyrene at low defined oxygen concentrations by a Mycobacterium sp.

In a fermentor, a Mycobacterium sp. was grown on pyrene at defined oxygen concentrations in a range from 11.4 to 227 microM. The maximal growth rate (mumax = 0.057 h-1) and the dissolved oxygen half-saturation constant (KDO = 5.9 microM) were calculated. At 3.4 microM, the growth rate (mu = 0.011 h-1) was only half of what was expected from the kinetic data. Apparently, this was due to limitation of an oxygenase of pyrene degradation.

Biodegradation, Environmental↗
PubMed1997

Novel metabolites in phenanthrene and pyrene transformation by Aspergillus niger.

Aspergillus niger, isolated from hydrocarbon-contaminated soil, was examined for its potential to degrade phenanthrene and pyrene. Two novel metabolites, 1-methoxyphenanthrene and 1-methoxypyrene, were identified by conventional chemical techniques. Minor metabolites identified were 1- and 2-phenanthrol and 1-pyrenol. No 14CO2 evolution was observed in either [14C]phenanthrene or [14C]pyrene cultures.

Alkaloids↗
PubMed2003

Degradation of benzo[a]pyrene by the litter-decomposing basidiomycete Stropharia coronilla: role of manganese peroxidase.

The litter-decomposing basidiomycete Stropharia coronilla, which preferably colonizes grasslands, was found to be capable of metabolizing and mineralizing benzo[a]pyrene (BaP) in liquid culture. Manganese(II) ions (Mn(2+)) supplied at a concentration of 200 micro M stimulated considerably both the conversion and the mineralization of BaP; the fungus metabolized and mineralized about four and twelve times, respectively, more of the BaP in the presence of supplemental Mn(2+) than in the basal medium. This stimulating effect could be attributed to the ligninolytic enzyme manganese peroxidase (MnP), whose activity increased after the addition of Mn(2+). Crude and purified MnP from S. coronilla oxidized BaP efficiently in a cell-free reaction mixture (in vitro), a process which was enhanced by the surfactant Tween 80. Thus, 100 mg of BaP liter(-1) was converted in an in vitro reaction solution containing 1 U of MnP ml(-1) within 24 h. A clear indication was found that BaP-1,6-quinone was formed as a transient metabolite, which disappeared over the further course of the reaction. The treatment of a mixture of 16 different polycyclic aromatic hydrocarbons (PAHs) selected by the U.S. Environmental Protection Agency as model standards for PAH analysis (total concentration, 320 mg liter(-1)) with MnP resulted in concentration decreases of 10 to 100% for the individual compounds, and again the stimulating effect of Tween 80 was observed. Probably due to their lower ionization potentials, poorly bioavailable, high-molecular-mass PAHs such as BaP, benzo(g,h,i)perylene, and indeno(1,2,3-c,d)pyrene were converted to larger extents than low-molecular-mass ones (e.g., phenanthrene and fluoranthene).

Basidiomycota↗
PubMed1997

Benzo(a)pyrene-albumin adducts in humans exposed to polycyclic aromatic hydrocarbons in an industrial area of Poland.

OBJECTIVES: The interaction of benzo(a)pyrene with serum albumin was measured in an attempt to identify the actual exposure and to evaluate albumin adduct measurements as biomarkers for exposure monitoring. METHODS: Benzo(a)pyrene-diol-epoxide (BPDE)-albumin adducts were measured by competitive enzyme linked immunosorbent assay (ELISA) in plasma of coke oven plant workers from three plants and from people living in a highly industrialised area of Silesia in Poland. Due to the high air concentrations of polycyclic aromatic hydrocarbons (PAHs) in this area, a control group was selected from a rural non-industrialised area in Poland. Breathing zone air measurements of PAHs were collected from some of the participants. RESULTS: Coke oven plant workers and non-occupationally exposed people had similar concentrations of albumin adducts whereas the rural controls were significantly lower (2.74 fmol adducts/microgram albumin (SEM 0.124)). The mean concentration of BPDE-albumin adduct in plasma of both the occupational and the environmental groups were significantly higher in the summer samples (4.34 fmol adducts/microgram albumin (SEM 0.335) and 4.55 fmol adducts/microgram albumin (SEM 0.296), respectively) than in the winter samples (3.06 fmol adducts/microgram albumin (SEM 0.187) and 3.04 fmol adducts/microgram albumin (SEM 0.184), respectively) even though the air measurements showed higher concentrations of PAHs in the winter. The statistical analysis did not show any effects of air exposures on concentrations of BPDE-albumin adduct. CONCLUSIONS: A multiple regression analysis of the measured concentrations of BPDE-albumin adducts for all the groups, during both seasons, indicates that occupational exposures do not contribute significantly to the formation of adducts. In general, the concentrations of albumin adducts found vary within relatively small limits for the two seasons and between the various groups of participants. No extreme differences were found.

Adolescent↗
PubMed2006

Black raspberry extracts inhibit benzo(a)pyrene diol-epoxide-induced activator protein 1 activation and VEGF transcription by targeting the phosphotidylinositol 3-kinase/Akt pathway.

Previous studies have shown that freeze-dried black raspberry extract fractions inhibit benzo(a)pyrene [B(a)P]-induced transformation of Syrian hamster embryo cells and benzo(a)pyrene diol-epoxide [B(a)PDE]-induced activator protein-1 (AP-1) activity in mouse epidermal Cl 41 cells. The phosphotidylinositol 3-kinase (PI-3K)/Akt pathway is critical for B(a)PDE-induced AP-1 activation in mouse epidermal Cl 41 cells. In the present study, we determined the potential involvement of PI-3K and its downstream kinases on the inhibition of AP-1 activation by black raspberry fractions, RO-FOO3, RO-FOO4, RO-ME, and RO-DM. In addition, we investigated the effects of these fractions on the expression of the AP-1 target genes, vascular endothelial growth factor (VEGF) and inducible nitric oxide synthase (iNOS). Pretreatment of Cl 41 cells with fractions RO-F003 and RO-ME reduced activation of AP-1 and the expression of VEGF, but not iNOS. In contrast, fractions RO-F004 and RO-DM had no effect on AP-1 activation or the expression of either VEGF or iNOS. Consistent with inhibition of AP-1 activation, the RO-ME fraction markedly inhibited activation of PI-3K, Akt, and p70 S6 kinase (p70(S6k)). In addition, overexpression of the dominant negative PI-3K mutant delta p85 reduced the induction of VEGF by B(a)PDE. It is likely that the inhibitory effects of fractions RO-FOO3 and RO-ME on B(a)PDE-induced AP-1 activation and VEGF expression are mediated by inhibition of the PI-3K/Akt pathway. In view of the important roles of AP-1 and VEGF in tumor development, one mechanism for the chemopreventive activity of black raspberries may be inhibition of the PI-3K/Akt/AP-1/VEGF pathway.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed2005

DNA damage from polycyclic aromatic hydrocarbons measured by benzo[a]pyrene-DNA adducts in mothers and newborns from Northern Manhattan, the World Trade Center Area, Poland, and China.

Polycyclic aromatic hydrocarbons (PAH), of which benzo[a]pyrene is a representative member, are combustion-related environmental pollutants and include known carcinogens. Laboratory animal studies indicate that the dose of PAHs to the fetus is on the order of a 10th that to the mother and that there is heightened susceptibility to PAH-induced carcinogenesis during the fetal and infancy periods. Carcinogen-DNA adducts, a measure of procarcinogenic genetic damage, are considered a biomarker of increased cancer risk. Here we compare the levels of benzo[a]pyrene-DNA adducts as a proxy for PAH-DNA damage measured in maternal blood and newborn cord blood obtained at delivery in four different populations of mothers (total of 867) and newborns (total of 822), representing a 30-fold range of exposure to ambient PAHs. The populations include residents in Northern Manhattan, participants in a study of the effects of the World Trade Center disaster, residents in Krakow, Poland, and residents in Tongliang, China. Mean adduct concentrations in both maternal and cord blood and the proportion of samples with detectable adducts, increased across the populations [Northern Manhattan < World Trade Center (WTC) < Krakow < Tongliang], consistent with the trend in estimated ambient exposure to PAHs (P < 0.001). For mothers, the means in the respective populations were Northern Manhattan (0.21 adducts per 10(8) nucleotides), WTC (0.23 adducts per 10(8) nucleotides), Krakow (0.28 adducts per 10(8) nucleotides), Tongliang (0.31 adducts per 10(8) nucleotides); the corresponding means in the newborns were Northern Manhattan (0.23), WTC (0.24), Krakow (0.29), Tongliang (0.31). The percentage of mothers with detectable levels of adducts in the respective populations were Northern Manhattan (36.8%), WTC (57.5%), Krakow (72.9%), Tongliang (73.4%); the corresponding percentages among the newborns were Northern Manhattan (42.4%), WTC (60.6%), Krakow (71.1%), Tongliang (79.5%). Despite the estimated 10-fold lower PAH dose to the fetus based on laboratory animal experiments, the adduct levels in the newborns were similar to or higher than in the mothers. This study suggests that the fetus may be 10-fold more susceptible to DNA damage than the mother and that in utero exposure to polycyclic aromatic hydrocarbons may disproportionately increase carcinogenic risk. The data support preventive policies to limit PAH exposure to pregnant women and children.

Adult↗
PubMed1993

Benzo(a)pyrene enhances atherosclerosis in White Carneau and Show Racer pigeons.

Benzo(a)pyrene (BaP), a major environmental pollutant and component of cigarette smoke, is both carcinogenic and atherogenic in experimental models. We investigated the effect of long-term administration of BaP on atherogenesis in both atherosclerosis-susceptible White Carneau (WC) and atherosclerosis-resistant Show Racer (SR) pigeons. The number and size of arterial lesions in the brachiocephalic arteries in WC and SR females but not males were significantly enhanced after long-term dosing with BaP. Metabolic activation appears to be required for BaP atherogenicity, since benzo(e)pyrene (BeP), a noncarcinogenic analogue of BaP, did not enhance lesion development. Studies with 3H-BaP revealed no significant differences between male and female or between WC and SR pigeons in the arterial distribution of BaP and/or its metabolites. There were no consistent differences in blood pressure or plasma cholesterol levels between breeds or sexes. However, chronic administration of BaP did result in complete infertility in female birds, concomitant with grossly visible changes in ovarian appearance. These results clearly show that long-term dosing with BaP alters ovarian structure and function in treated birds, at the same time aggravating the development of arterial lesions. Thus, BaP-induced atherogenicity in female pigeons may be a consequence of an alteration in estrogen production or of antiestrogenic properties of BaP at the level of the arterial wall and may serve as a highly useful animal model to examine the well-known rapid development of atherosclerosis in postmenopausal women.

Animals↗
PubMed2002

Lung-specific expression of dominant-negative mutant p53 in transgenic mice increases spontaneous and benzo(a)pyrene-induced lung cancer.

Mutations in the p53 gene have been implicated to play an important role in the development of various human cancers. To evaluate the importance of p53 in lung cancer, a transgenic mouse model was established by utilizing the Clara cell secretory protein (CCSP) promoter to target the expression of a dominant-negative mutant form of p53 (dnp53) in the lung. In two transgenic CCSP-dnp53 founder lines, the dnp53 protein was expressed exclusively in the lungs. The incidence of spontaneous lung cancer in 18-month-old transgenic mice was 45%, whereas that in age-matched control mice was 20%. The relative risk of lung tumors in CCSP-dnp53 mice was 2.3 times that of wild-type mice (exact confidence limits of 0.69, 17.5). In addition to the increased incidence of spontaneous lung tumor, these mice were more susceptible to the development of lung adenocarcinoma after exposure to benzo(a)pyrene (BaP). Six months after intratracheal instillation of benzo(a)pyrene, the tumor incidence in wild-type and CCSP-dnp53 mice was 39% and 73%, respectively. The risk of lung tumors was 25.3 times greater in BaP-treated mice adjusted for transgene expression (95% confidence limits of 3.29, 678, mid-p corrected). These results suggest that p53 function is important for protecting mice from both spontaneous and BaP-induced lung cancers.

Adenocarcinoma↗
PubMedDate not supplied

Nanoparticles from polylactide and polyether block copolymers: formation, properties, encapsulation, and release of pyrene--fluorescent model of hydrophobic drug.

Polylactide-b-polyglycidol-b-poly(ethylene oxide) terpolymers and their derivatives with carboxyl and 4-(phenylazo)phenyl labels in polyglycidol blocks were used for formation of nanoparticles. Nanoparticles were produced by self assembly of terpolymer macromolecules in water above the critical aggregation concentration and by dialysis of terpolymer solutions in 1,4-dioxane against water. For terpolymers with 4-(phenylazo)phenyl labels critical aggregation concentrations increased after irradiation with UV light (300 < lambda < 400 nm) inducing conformational change of the label from trans- to cis-conformation. Diameters of nanoparticles obtained by self-assembly of macromolecules ranged from 20 to 44 nm. Dialysis yielded nanoparticles with bimodal diameter distribution. One fraction had diameters below 35 nm and diameters of the second fraction were in a range from 350 to 2300 nm, depending on terpolymer structure. Mixtures of terpolymers with poly(L,L-lactide) and poly(D,D-lactide) blocks yielded nanoparticles with diameters from 350 to 440 nm. Pyrene was incorporated into nanoparticles by partition between solution and nanoparticles or directly during particle formation by dialysis. Monitoring of pyrene release from nanoparticles suggests that a fraction of this compound was entrapped into the polylactide core whereas the remaining one was located in the polyether rich shell. The release from shells is faster for nanoparticles made from copolymers with carboxyl labels in polyglycidol blocks.

Capsules↗
PubMed1984

Fluorescence measurements of DNA-bound metabolites of benzo(a)pyrene derivatives with different carcinogenic effects.

(+/-)-trans-dihydroxy-7,8-dihydrobenzo(a)pyrene (BP-7,8-diol) and 9-hydroxybenzo(a)pyrene (9-OH-BP) were metabolized by rat liver microsomes in the presence of calf thymus DNA, resulting in preferential DNA binding of fluorescent (+)-anti-BP-7,8-diol-9,10-epoxide (BPDE) and 9-OH-BP-4,5-epoxide, respectively. When the DNA is denatured the fluorescence intensities of the bound metabolites change in a characteristic manner. Fluorescence decay measurements show that the intensity changes are due to changes in lifetimes of the excited states. Model substances for the bound metabolites were studied in solvents of different polarity. We found that the fluorescence changes observed after denaturation of the DNA may be explained as solvent polarity effects, so that denaturation forces the bound metabolites from a more hydrophobic environment to a hydrophilic one. Fluorescence depolarization studies as a function of temperature in combination with previous linear dichroism studies show that both BPDE and 9-OH-BP-4,5-epoxide form rigidly associated complexes with native DNA.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1997

Detection of benzo[a]pyrene-DNA adducts in human placenta and umbilical cord blood.

Placenta constitutes a vital organ of exchange between mother and foetus. In addition to this favourable effect for foetal development, placenta indirectly may allow transfer of several maternal blood xenobiotics. Human placenta and umbilical cord blood are interesting models for investigating maternal environment and the metabolism, the bioactivation and the transfer of carcinogens such as polycyclic aromatic hydrocarbons. We used them to assess the effect of a woman's smoking on the foetus. Few studies cover this subject. In pregnant women who have continued to smoke, benzo[a]pyrene compound of cigarette smoke is metabolically activated to diol-epoxide derivative: benzo[a]pyrene-trans-7,8-dihydrodiol-9,10-epoxide, ultimate carcinogen (BPDE-I). This derivative is covalently fixed on DNA and gives BPDE-I-DNA adducts. By a competitive immunoassay technique, we determined BDPE-I-DNA adducts in 20 samples of placenta and umbilical cord blood from women who smoked (n = 15) and who did not (n = 10). Tobacco consumption was checked by urinary cotinine determination. In the group of smokers levels of adducts were found in 13 specimens of placenta (from 10 to 60 fmol/50 micrograms of DNA) and 12 umbilical cord blood (from 10 to 22.15 fmol/50 micrograms of DNA) samples. These results indicate that a mother's tobacco consumption is linked to the accumulation of BPDE-I-DNA adducts in the placenta, which are seen in smaller quantities in the umbilical cord blood, probably because of the metabolic capacity of the placenta and the transfer of B[a]P from the mother to the foetus.

Adolescent↗
PubMed2006

Time- and concentration-dependent changes in gene expression induced by benzo(a)pyrene in two human cell lines, MCF-7 and HepG2.

BACKGROUND: The multi-step process of carcinogenesis can be more fully understood by characterizing gene expression changes induced in cells by carcinogens. In this study, expression microarrays were used to monitor the activity of 18,224 cDNA clones in MCF-7 and HepG2 cells exposed to the carcinogen benzo(a)pyrene (BaP) or its non-carcinogenic isomer benzo(e)pyrene (BeP). Time and concentration gene expression effects of BaP exposure have been assessed and linked to other measures of cellular stress to aid in the identification of novel genes/pathways involved in the cellular response to genotoxic carcinogens. RESULTS: BaP (0.25-5.0 muM; 6-48 h exposure) modulated 202 clones in MCF-7 cells and 127 in HepG2 cells, including 27 that were altered in both. In contrast, BeP did not induce consistent gene expression changes at the same concentrations. Significant time- and concentration-dependent responses to BaP were seen in both cell lines. Expression changes observed in both cell lines included genes involved in xenobiotic metabolism (e.g., CYP1B1, NQO1, MGST1, AKR1C1, AKR1C3,CPM), cell cycle regulation (e.g., CDKN1A), apoptosis/anti-apoptosis (e.g., BAX, IER3), chromatin assembly (e.g., histone genes), and oxidative stress response (e.g., TXNRD1). RTqPCR was used to validate microarray data. Phenotypic anchoring of the expression data to DNA adduct levels detected by 32P-postlabelling, cell cycle data and p53 protein expression identified a number of genes that are linked to these biological outcomes, thereby strengthening the identification of target genes. The overall response to BaP consisted of up-regulation of tumour suppressor genes and down-regulation of oncogenes promoting cell cycle arrest and apoptosis. Anti-apoptotic signalling that may increase cell survival and promote tumourigenesis was also evident. CONCLUSION: This study has further characterised the gene expression response of human cells after genotoxic insult, induced after exposure to concentrations of BaP that result in minimal cytotoxicity. We have demonstrated that investigating the time and concentration effect of a carcinogen on gene expression related to other biological end-points gives greater insight into cellular responses to such compounds and strengthens the identification of target genes.

Benzo(a)pyrene↗
PubMed1991

[Fluorescence label of alcohols and amines with pyrene-1-carbonyl fluoride].

Pyrene-1-carbonyl fluoride (PCF) was synthesized as a precolumn fluorescent labeling reagent for alcohols and amines for use in high performance liquid chromatography (HPLC). PCF reacted with primary and phenolic hydroxyl groups in dichloromethane at 100 degrees C for 30 min in the presence of 4-dimethylaminopyridine (DMAP) to give the corresponding fluorescent pyrene esters and also reacted with primary amines in acetonitrile at room temperature for 2 min in the presence of DMAP to give the amides. The PCF esters of corticosteroids were separated by normal-phase chromatography on a Cosmosil 5 SL column with hexane-ethyl acetate (6:5, v/v) and the PCF amides of primary amines were separated by reversed-phase chromatography on a TSK gel ODS-80 TM column with methanol-water (10:3, v/v). The detection limits (S/N = 3) of cortisone and 2-phenylethylamine were 600 fmol and 800 fmol for an injection volume of 10 microliters, respectively.

Alcohols↗
PubMed1994

Effects of induction and age-dependent enzyme expression on lung bioavailability, metabolism, and DNA binding of urban air particulate-absorbed benzo[a]pyrene, 2-nitrofluorene, and 3-amino-1,4-dimethyl-5H-pyridol-(4,3)-indole.

The effect of interactions between urban air particulates (UAP) and carcinogens on bioavailability, metabolism, and DNA binding was studied in the isolated perfused and ventilated rat lung. The availability of benzo[a]pyrene (B[a]P) varied from 29 to 60% after intratracheal doses of carcinogen particulates dissolving extremely slow and fast, respectively. Several cytochrome P450 enzyme (P450) inducers acting as 2,3,7,8-tetrachlorodibenzo[p]dioxin-receptor ligands have been identified in UAP extracts. beta-Naphthoflavone (BNF) was used to study how P450 induction alters the lung metabolism of carcinogens. Pretreatment increased the lung clearance for B[a]P 8-fold and for 2-nitrofluorene (2NF) by a factor of four from 0.55 +/- 0.06 ml/min to 2.37 +/- 0.62 ml/min. Studies with the intact lung and with isolated lung cells show that carcinogen metabolism and pharmacokinetics depend both on the route of exposure and dosage and on the distribution of specific enzymes. A cytochrome P450IIB1 enzyme was detected in lung epithelial cells where it catalyzes 9-hydroxylation of 2NF. This rat lung 2NF-9-hydroxylation capacity increases in parallel with the age dependent up-regulation of lung P450IIB1 expression. Both human and rat lung tissue have the capacity to form 9-hydroxy-2-nitrofluorene (9-OH-2NF) that is mutagenic. A BNF-inducible P450IA1 was detected in endothelial and alveolar type II cells. Consequently, aromatic hydroxylation dominated when 2NF was dosed directly into the lung circulation. Pretreatment of rats with BNF before intratracheal B[a]P dosage induced lung cytochrome P450IA1. The 7,8-dihydroxy-9,10-oxybenzo[a]pyrene-deoxyguanosine adduct and the total lung DNA adduct levels increased significantly from a peak level of 75 +/- 8 to 151 +/- 19 fmole/mg DNA in lungs from control and BNF pretreated rats, respectively. 3-Amino-1,4-dimethyl-5H-pyridol-(4,3)-indole (TRP-P1) is a potent mutagen and carcinogen identified in UAP extracts. Dietary BNF pretreatment of rats altered the [14C]TRP-P1 distribution as analyzed by whole-body autoradiography. An enhanced retention was observed in the small intestine, forestomach, esophagus, and lung. UAP catalyzes oxygen radical formation and deoxyguanosine-8-hydroxylation, which were inhibited when the UAP samples were extracted with organic solvents or when they were incubated in the presence of desferroxamine. We therefore postulate that a polycyclic aromatic hydrocarbon autooxidation pathway may be responsible for generation of hydrogen peroxide, which may be further converted to hydroxyl radicals through an iron-dependent reaction.

Aging↗
PubMed1985

Quantitation of exposure to benzo[a]pyrene with monoclonal antibodies.

It is now possible to quantitate carcinogen adducts on DNA by highly sensitive immunoassays. These techniques are particularly useful for screening human populations for exposure to potential environmental carcinogens. We have developed a panel of monoclonal antibodies that react with benzo(a)pyrene (BP) modified DNA to be used in an enzyme linked immunoassay (ELISA) to quantitate adduct levels of both human and animal samples. BALBc/Cr mice were immunized with either DNA modified by 7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9, 10-tetrahydrobenzo(a)pyrene (BPDE-I-DNA) complexed electrostatically to methylated bovine serum albumin or with BPDE-I-modified guanosine conjugated with bovine serum albumin (BPDE-I-G-BSA). Four stable clones were produced from the spleen cells of animals immunized with BPDE-I-DNA and one from BPDE-I-G-BSA immunized animals. All antibodies were shown to be highly specific for BPDE-I-DNA and did not crossreact with nonmodified DNA or with N-2-acetylaminofluorene or 1-aminopyrene modified DNA. The antibodies differed in their sensitivity to BPDE-II-DNA, BPDE-I-poly G, BPDE-I-tetraols and BPDE-I-dG. In general, all the antibodies showed the greatest affinity for their original antigen. Those generated against modified DNA showed highest reactivity against modified DNA while the one antibody generated against the monoadduct showed highest reactivity with the monoadduct. These antibodies are currently being used in a highly sensitive competitive ELISA to quantitate levels of BP-DNA adducts in various animal and human tissue samples.

Animals↗
PubMed1997

Neutrophils amplify the formation of DNA adducts by benzo[a]pyrene in lung target cells.

Inflammatory cells and their reactive oxygen metabolites can cause mutagenic effects in lung cells. The purpose of this study was to investigate the ability of activated neutrophils to modulate DNA binding of benzo[a]pyrene (B[a]P), a known carcinogen, in lung target cells. Equivalent numbers of rat lung epithelial cells (RLE-6TN cell line) and freshly isolated human blood neutrophils (PMN) were coincubated in vitro for 2 hr after addition of benzo[a]pyrene (0.5 microM) or two of its trans-diol metabolites, with or without stimulation with phorbol myristate acetate (PMA). DNA adducts of B[a]P-metabolites were determined in target cells using 32P-postlabeling; oxidative DNA damage (7-hydro-8-oxo-2'-deoxyguanosine [8-oxodG]) was evaluated by high performance liquid chromatography with electrochemical detection. Increased DNA adducts were observed in lung cells coincubated with polymorphonuclear leukocytes (PMN). Activation of PMN with PMA, or addition of more activated PMN in relation to the number of lung cells, further increased the number of adducts, the latter in a dose-response manner. Incubation with B[a]P-4,5-diol did not result in any adduct formation, while B[a]P-7,8-diol led to a significant number of adducts. Moreover, PMA-activated PMN strongly enhanced adduct formation by B[a]P-7,8-diol, but not 8-oxodG, in lung cells. The addition of antioxidants to the coincubations significantly reduced the number of adducts. Results suggest that an inflammatory response in the lung may increase the biologically effective dose of polycyclic aromatic hydrocarbons (PAHs), and may be relevant to data interpretation and risk assessment of PAH-containing particulates.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed2006

Differential detection of phospholipid fluidity, order, and spacing by fluorescence spectroscopy of bis-pyrene, prodan, nystatin, and merocyanine 540.

The properties of liquid-ordered, solid-ordered, and liquid-disordered phases were investigated by steady-state fluorescence spectroscopy in liposomes composed of mixtures of dipalmitoylphosphatidylcholine and cholesterol (0-40 mol %) as a function of temperature (24-51 degrees C). The fluorescent probes used (bis-pyrene, nystatin, prodan, and merocyanine) were chosen because they differ in the location they occupy in the membrane and in the types of properties they sense. Comparison of phase diagrams with contour plots of the fluorescence data suggested that bis-pyrene is sensitive primarily to lipid order. In contrast, nystatin fluorescence intensity responded to changes in lipid fluidity. The shape of the prodan emission spectrum detected both liquid-solid and order-disorder transitions in the phase diagram. Merocyanine's behavior was more complex. First, it was more sensitive than any of the other probes to the membrane pretransition that occurs in the absence of cholesterol. Second, regardless of whether emission intensity, anisotropy, or spectral shape was observed, the probe appeared to distinguish two types of liquid-ordered phases, one with tightly packed lipids and one in which the apparent spacing among lipids was increased. The prodan data supported these results by displaying modest versions of these two observations. Together, the results identify eight regions within the phase diagram of distinguishable combinations of these physical properties. As an example of how this combined analysis can be applied to biological membranes, human erythrocytes were treated similarly. Temperature variation at constant cholesterol content revealed three of the eight combinations identified in our analysis of liposomes.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2005

Stage change in binding of pyrene to selected humic substances under different ionic strengths.

Influence of ionic strength (I) on the pyrene binding constant (Koc) in six different humic substance (HS) solutions, three soil humic acids, one freshwater humic acid, one freshwater fulvic acid, and one freshwater natural organic matter was investigated by fluorescence quenching technique. Specific two-stage changes of Koc with increasing ionic strength were only observed in three soil humic acid solutions with varying to different extents. Two primary governing factors for the differences of these HS systems are binding potential and the abundance of acidic functional groups (carboxyl and phenolic) of HS. Experimental results demonstrated that within the ionic strength range studied in the present research, ionic strength had a major effect on molecular size of HS and brought about change of Koc. The salting-out effect did not. The change in molecular size was caused by the associations between cations and acidic groups of HS. Using pyrene as a probe, we also investigated the interaction of cation and dissolved HS and found relatively slow kinetics (at least 5 d), leading us to believe that the special cation-HS kinetics may influence the results of geochemical studies involving the measurement of ionic strength effect.

Adsorption↗