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Biochemical characterization of the interactions of the novel pleuromutilin derivative retapamulin with bacterial ribosomes.

Retapamulin is a semisynthetic pleuromutilin derivative being developed as a topical antibiotic for treating bacterial infections of the skin. It is potent in vitro against susceptible and multidrug-resistant organisms commonly associated with bacterial skin infections. We report detailed mode of action studies demonstrating that retapamulin binds to the bacterial ribosome with high affinity, inhibits ribosomal peptidyl transferase activity, and partially inhibits the binding of the initiator tRNA substrate to the ribosomal P-site. Taken together, these data distinguish the mode of action of retapamulin from that of other classes of antibiotics. This unique mode of action may explain the lack of clinically relevant, target-specific cross-resistance of retapamulin with antibacterials in current use.

Animals↗

Natural mediators in the oxidation of polycyclic aromatic hydrocarbons by laccase mediator systems.

The oxidation of polycyclic aromatic compounds was studied in systems consisting of laccase from Trametes versicolor and so-called mediator compounds. The enzymatic oxidation of acenaphthene, acenaphthylene, anthracene, and fluorene was mediated by various laccase substrates (phenols and aromatic amines) or compounds produced and secreted by white rot fungi. The best natural mediators, such as phenol, aniline, 4-hydroxybenzoic acid, and 4-hydroxybenzyl alcohol were as efficient as the previously described synthetic compounds ABTS [2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)] and 1-hydroxybenzotriazole. The oxidation efficiency increased proportionally with the redox potentials of the phenolic mediators up to a maximum value of 0.9 V and decreased thereafter with redox potentials exceeding this value. Natural compounds such as methionine, cysteine, and reduced glutathione, containing sulfhydryl groups, were also active as mediator compounds.

Amines↗

[Measurement of various polyaromatic hydrocarbons in the urban area of Naples].

In order to investigate the organic compound fraction of the Naples aerosol a chromatographic method was used for the separation and analysis of the polycyclic aromatic (PAH). As a first step a suitable one-step thin-layer chromatography (TLC) separation of the cyclohexane extractable material from airborne particulate was sought. After the TLC separation the concentrated samples were analyzed by reverse-phase liquid chromatography with fluorescence detection. We obtained chromatographic separation of five PAH on the EPA Priority Pollutant List and we determined the concentration of these PAHs present in atmospheric matter.

Air Pollutants↗

Bitumen fumes: review of work on the potential risk to workers and the present knowledge on its origin.

Bitumens fumes contain polycyclic aromatic compounds (PAC). There is a possibility of long-term health effects following chronic exposure by inhalation or skin contamination in asphalt road pavers and highway maintenance workers. Epidemiological and experimental studies on this topic are reviewed and the possible causes of cancer discussed with a primary focus on heterocyclic polyaromatic compounds. In 2001, the results of the IARC epidemiological study confirmed an excess of lung cancer despite a lower cancer mortality. In vitro genotoxicity and mechanistic studies demonstrated a mutagenic effect of bitumen fume condensates (BFC) and some data suggested that the polycyclic aromatic hydrocarbons (PAH) analysed were not the major genotoxic compounds in bitumen fume condensates. Other compounds such as nitrogen-, sulfur- and/or oxygen-containing PAH or their alkyl substituted analogues, mutagenic in the Ames mutation assay, may be involved in the genotoxic effect of BFC. After skin painting with BFC, DNA adducts were found in skin, lung and lymphocytes of all the treated animals. Differences in the adduct patterns were also observed, but a more polar adduct was common to the three tissues and not observed in those from rats treated with coal-tar fume condensates (CTFC). Rat inhalation experiments with bitumen fumes confirmed the presence of a DNA-adduct in the lungs with the same Rf as the previous polar adduct. This adduct therefore merits further investigation as a potential biomarker in lymphocyte DNA to follow exposed workers. All the analytical data and the mechanistic data are complementary and suggest the potential role of thiophenes in the genotoxicity of bitumen fumes. Some thiophenes have lower mutagenic activity than their isosteric PAH, whereas others are very potent carcinogens. Generally, the sulfur analogues of PAH (SPAH) in bitumen fumes have a higher concentration than the PAH of similar molecular weight, whereas the SPAH in coal-tar fumes have a much lower concentration than the corresponding PAH. This may explain why the more polar adducts have been detected only in animals exposed to bitumen fume. In a skin carcinogenicity study of condensed asphalt roofing fumes, it has been demonstrated that the most active fractions were those containing a variety of aromatic SPAH. In conclusion to this review, there is an interest in determining the chemical identity of the major DNA adducts induced by BFC. This would allow experimental studies on the carcinogenic potency of these compounds and their validation as potential biomarkers. These compounds could thus merit further analytical investigation in preference to the PAH included in the list of the US Environmental Protection Agency that are currently being analysed by the industry in field studies.

Administration, Cutaneous↗

Pilot survey of a broad range of priority pollutants in sediment and fish from the Ebro river basin (NE Spain).

Priority organic pollutants were investigated in sediments and fish collected along the Ebro river basin (NE Spain) to evaluate their occurrence, transport and bioavailability. Sediments were collected in 18 sites and two species of fish were captured in nine sites according to the availability in each area. The sampling sites covered industrial, urban and agricultural areas. Four methods were used to detect 20 organochlorine compounds (OCs), 8 polycyclic aromatic hydrocarbons, 3 organotin compounds, 2 alkylphenols and 40 polybrominated diphenyl ethers (PBDEs) from purified extracts. The contamination pattern was site specific and no downstream increase in concentration of pollutants was observed but rather a generalized low level diffuse pollution. Target compounds were detected in sediments at 0.01 to 2331 microg/kg dry weight, and only OCs and PBDEs were accumulated in benthopelagic fish. Toxicological assessment was performed according to predicted environmental levels and revealed sites where adverse effects could occur.

Animals↗

Polyvinylpyrrolidone-coated silica packings for chromatography of proteins and peptides.

Polyvinylpyrrolidone (PVP)-coated silica sorbents were synthesized by interaction of a copolymer of vinyl-methyldiethoxysilane and vinylpyrrolidone with LiChrospher Si 300 and LiChrospher Si 500 silicas. The coating procedure retains the wide-pore structure of the starting silicas, as was shown by nitrogen adsorption, mercury porosimetry and inverse size-exclusion chromatographic measurements. Good selectivity and separation ability of the synthesized packings toward proteins and peptides were demonstrated in the hydrophobic interaction chromatographic mode of separation. Aromatic compounds undergo a specific interaction with bonded PVP chains which can be used for the preconcentration or selective recovery of polycyclic aromatic hydrocarbons.

Amino Acid Sequence↗

Lignan compounds and 4,4'-dihydroxybiphenyl protect C2C12 cells against damage from oxidative stress.

Lignan compounds are known to have various biological activities, especially antioxidative effects. We investigated whether lignan compounds show antioxidative activity in myoblast C2C12 cells. Among 14 lignan compounds investigated, two lignans containing two phenolic functional groups, namely Gomisin J and GR-12, prevented hydrogen peroxide (H(2)O(2))-induced cell death. A simple compound, 4,4'-dihydroxybiphenyl, which was found to be a common component of Gomisin J and GR-12, also largely prevented H(2)O(2)-induced cell death and almost completely prevented H(2)O(2)-induced increases in p38 MAPK phosphorylation. Our present results provide a useful in vitro system for clarifying the molecular mechanisms of lignan-mediated antioxidative effects and evaluating lead molecules toward the development of therapeutic drugs.

Animals↗

Mutation spectra of chemical mutagens determined by Lac+ reversion assay with Escherichia coli WP3101P-WP3106P tester strains.

We previously reported the development of mutation-specific Escherichia coli B tester strains WP3101 to WP3106 from strain WP2uvrA. In this study we constructed their pKM101-containing derivatives WP3101P to WP3106P, and further isolated their rfa derivatives WP4101-WP4106 and WP4101P-WP4106P. The six kinds of F' plasmids (lacI-, lacZ-, proAB+), each of which carries a different lacZ allele, contained in the above strains were originally derived from E. coli K-12 strains CC101-CC106. All the tester strains show Lac- and Trp- phenotype. Assays for transitions and transversions are based upon Lac+ reversion of a specific mutation located within the lacZ gene on an F' plasmid. The trpE65(ochre) allele in the same strains enables them to be used for Trp+ reversion assays as well. In the present paper, we evaluated the sensitivity, specificity, and usefulness of the newly developed tester strains. Strains WP3101P-WP3106P were highly sensitive to determine mutational profile of heterocyclic amines with S9 mix-mediated metabolic activation and most of the oxidative mutagens and free radical generators tested. Every type of base-pair substitutions induced by 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) or 5-diazouracil were detected in strains WP3101P-WP3106P, while A:T-->C:G and G:C-->A:T mutations induced by MeIQ, and A:T-->C:G, G:C-->A:T, and G:C-->C:G by 5-diazouracil were not detected in pKM101-free tester strains. In pKM101-carrying strains, cumene hydroperoxide induced all types of base substitutions, while formaldehyde preferentially induced G:C-->T:A transversions. Phenazine methosulfate induced predominantly G:C-->A:T transitions and G:C-->T:A transversions, while H2O2 induced predominantly G:C-->T:A and A:T-->T:A transversions. Introduction of the rfa mutation considerably enhanced sensitivity to bulky mutagens such as polycyclic aromatic compounds. All six possible base substitutions induced by 9, 10-dimethyl-1,2-benzanthracene (DMBA) were detected in tester strains WP4101P-WP4106P. In conclusion, our tester strains WP3101P-WP3106P and WP4101P-WP4106P permitted rapid and simple detection of specific mutations induced by variety of mutagens.

Animals↗

One-step transformation of tricyclopentabenzene (trindane, C(15)H(18)) to bicyclo(10.3.0)pentadec-1(12)ene- 2,6,7,11-tetrone (C(15)H(18)O(4)) and its aldol product, 12-hydroxy-16-oxatetracyclo(10.3.1.0.(1,5)0(7,11))hexadec-7(11)ene-2,6-dione (C(15)H(18)O(4)).

[reaction: see text] Ozonolysis of 1 largely results in 2 and 3, having features similar to several classes of natural products. The retention of the C(15) pericycle suggests preference for the cleavage of pi-bonds endo to the cyclopentane ring. This unique property of trindane offers opportunities for synthesis of complex natural products from this hydrocarbon that can be made in quantity by acid-catalyzed trimerization of cyclopentanone.

Bridged Bicyclo Compounds↗

Covalent binding of components of coal-tar, creosote and bitumen to the DNA of the skin and lungs of mice following topical application.

In order to assess the DNA damaging ability of complex carcinogenic mixtures, male Parkes mice were treated topically with solutions of (i) pharmaceutical coal-tar, (ii) creosote, a blend of coal-tar fractions or (iii) bitumen, a product of oil-refining. DNA was isolated from the treated skin and analysed by 32P-post-labelling. A band of radioactivity was obtained on polyethyleneimine--cellulose TLC indicating the formation of DNA adducts by a large number of different chemical compounds present in these fossil fuel products. The chromatographic properties of the adducts were broadly characteristic of those formed by polycyclic aromatic hydrocarbons. The levels of DNA binding were approximately 0.4 fmol total adducts/micrograms DNA 24 h after treatment for coal-tar and creosote and 0.09 fmol/micrograms DNA for bitumen treatment. The persistence of adducts in mouse skin following a single dose of either coal-tar or creosote was found to exhibit a phase of rapid removal, in which one half to two thirds of the initial levels of adducts, detected at 24 h after treatment, were removed by 7 days followed by a second phase in which one half to two thirds of the remainder was removed in the succeeding 25 days. When mice were treated topically with multiple carcinogenic doses of coal-tar, creosote or bitumen for up to 5 weeks, a steady accumulation of adducts was seen in skin DNA during the course of the treatment, approaching a steady-state level towards the end of the treatment period in some instances. A similar accumulation of adducts was also evident in lung DNA, the levels being approximately half those attained in skin. The results demonstrate the application of 32P-post-labelling to the detection of DNA adducts formed in vivo by complex carcinogenic mixtures of the type to which humans are exposed.

Administration, Topical↗