Microbial diversification of Diels-Alder cycloadducts by whole cells of Penicillium brasilianum.

Functionalizations of cycloadducts are important steps for the use of Diels-Alder reactions in the construction of complex cyclic or polycyclic molecules from relatively simple starting materials. In the present work, we studied the ability of Penicillium brasilianum to perform microbial transformations of racemic Diels-Alder endo-cycloadducts. Thus, Diels-Alder products, obtained from reacting cyclopentadiene or 2,3-dimethylbutadiene with alkylated para-benzoquinones, were transformed by the resting cells of P. brasilianum producing new functionalized polycyclic compounds. These biotransformations yielded novel products of oxidation and ring closure, reduction of the C=C or C=O in [Formula: see text]-unsaturated system, and allylic hydroxylations. The reduction products (conjugated double bond and carbonyl group) were also synthesized, and the enantioselectivity of both in vitro and in vivo processes was evaluated. In all cases, the microbiological transformations were enantioselective. In silico docking studies of the Diels-Alder cycloadducts with P. brasilianum oxidoreductase "old yellow enzymes" shed more light on these transformations.

Predicting bioaccumulation of PAHs in the trophic chain in the estuary region of Paranagua, Brazil.

The presence of polycyclic aromatic hydrocarbon (PAH) compounds in sediment and water samples collected in the estuary area of Paranagua, southern Brazil, was investigated. There is a lot of port activity in the region. Recreational fishing is widespread; thus, there is concern about possible contamination by PAHs. The 16 priority PAHs were investigated, and only eight were found. The total concentration of PAHs ranged from 40.8 to 406.8 ng/g. High molecular weight were the most abundant, while PAHs with a low molecular weight were absent. There are suspicions that the main source of PAHs is combustion, but some uncertainties exist, and there may even be the presence of PAHs resulting from accidental spills of crude oil. Although the sediments contain PAHs, the amount is below the maximum concentrations allowed by the Brazilian environmental legislation, as well as the maximum levels at which adverse effects are observed. From the analytical results, a probable bioaccumulation was assessed in the local trophic chain using a mathematical model (Arnot and Gobas, Environ Toxicol Chem 23(10):2343-2355, 2004). The model showed that there is a possibility of biomagnification along the food chain selected. Three fishes with high local consumption were selected, and the concentration of some PAHs could be found in those fishes.

Bioconcentration factor estimates of polycyclic aromatic hydrocarbons in grains of corn plants cultivated in soils treated with sewage sludge.

This study presents a model to simulate the organic substance concentrations in corn grains assuming that the substances in soil solution are absorbed via the transpiration stream by plants growing in soils fertilized with sewage sludge (SS). The model was applied and validated using soil and corn grain samples from a long-term field experiment with six successive yearly applications of SS to the soil. The following polycyclic aromatic hydrocarbons (PAHs) were simulated and evaluated in soil and grain samples: acenaphthene, acenaphthylene, anthracene, benz(a)anthracene, benz(a)pyrene, benz(b)fluoranthene, benz(g,h,i)perylene, benz(k)fluoranthene, chrysene, dibenz(a,h)anthracene, fluoranthene, fluorene, indeno(1,2,3-c,d)pyrene, naphthalene, phenanthrene and pyrene. The PAH bioconcentration factors (BCF) in corn grains ranged from 1.57 to 10.97 L kg(-1). Polycyclic aromatic hydrocarbons with low soil distribution coefficients and high values of transpiration stream concentration factors (TSCF) are more likely to be absorbed by corn plants and accumulated in grains. It was possible to estimate and observe that highly lipophilic PAH molecules (heavy PAHs) show lower accumulative potential in corn grains than the less lipophilic ones (light PAHs). Sewage sludges containing significant concentrations of light PAHs with two, three or four benzene rings should be avoided as fertilizers in alimentary field crops.

Microbial consortium bioaugmentation of a polycyclic aromatic hydrocarbons contaminated soil.

In this study we evaluated the capacity of a defined microbial consortium (five bacteria: Mycobacterium fortuitum, Bacillus cereus, Microbacterium sp., Gordonia polyisoprenivorans, Microbacteriaceae bacterium, Naphthalene-utilizing bacterium; and a fungus identified as Fusarium oxysporum) isolated from a PAHs contaminated landfarm site to degrade and mineralize different concentrations (0, 250, 500 and 1000 mg kg(-1)) of anthracene, phenanthrene and pyrene in soil. PAHs degradation and mineralization was evaluated by gas chromatography and respirometry, respectively. The microbial consortium degraded on average, 99%, 99% and 96% of the different concentrations of anthracene, phenanthrene and pyrene in the soil, in 70 days, respectively. This consortium mineralized 78%, on average, of the different concentrations of the 3 PAHs in soil after 70 days. Contrarily, the autochthonous soil microbial population showed no substantial mineralization of the PAHs. Bacterial and fungal isolates from the consortium, when inoculated separately to the soil, were less effective in anthracene mineralization compared to the consortium. This signifies synergistic promotion of PAHs mineralization by mixtures of the monoculture isolates (the microbial consortium).

Determination of specific receptor sites for platelet activating factor in bovine neutrophils.

OBJECTIVE: To identify and characterize a platelet activating factor (PAF) receptor in bovine neutrophils by use of radioligand binding, reverse transcription-polymerase chain reaction (RT-PCR) assay, and western blot analysis. ANIMALS: 4 healthy adult cows. PROCEDURE: Bovine neutrophil membranes were isolated for association, dissociation, and saturation binding experiments with PAF labeled with hydrogen 3 (3H-PAF). The RT-PCR assay was performed with appropriate human primers, and western blot analysis was developed with a polyclonal antibody obtained from a peptide of bovine PAF receptor. RESULTS: Analysis of kinetic binding data supported a single class of PAF receptor. Binding of 3H-PAF to membrane preparations was selectively displaced by PAF and a nonhydrolyzable analogue of guanine triphosphate (Gpp[NH]p) and by lyso-PAF (a biologically inactive analogue of PAF) to a lesser extent. Among other PAF receptor antagonists, 14-deoxyandrographolide and WEB 2086 were the most effective in inhibiting 3H-PAF binding sites in neutrophil membranes; 2 lignans, schisandrin-A and gamma-schisandrin were also effective, but 2 gingkolides (BN52020 and BN52021) only mildly inhibited 3H-PAF binding. Results of RT-PCR assay and western blot analysis of neutrophil crude membranes confirmed the presence of a PAF receptor. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that bovine neutrophils express only 1 type of PAF receptor, and it is likely that this receptor is involved in inflammatory responses. The most effective PAF antagonists were 14-deoxyandrographolide and WEB 2086; these PAF antagonists may be potentially useful in the treatment of inflammatory processes in cattle.

Molecular mechanics explanation for the stereochemical and shape selectivity of B-DNA for "bay-region" carcinogens.

The equilibrium structures of 20 intercalated physical complexes of "bay-region" triol carbocations of polycyclic aromatic hydrocarbons (PAHs) with B-DNA are obtained by AMBER molecular modeling. The complexes with highly potent carcinogens are found (i) to undergo only minor conformational changes upon complexation, (ii) to be stabilized by hydrogen bonds between two hydroxyl groups of the triol carbocations and N3 atoms of the adjacent guanine residues, and (iii) to be "preorganized" for covalent bonding. A new explanation for the absolute stereochemical and shape dependence of carcinogenesis by PAHs is presented. The biologically active conformers of both carcinogenic stereoisomers (anti and syn) of triol carbocations are characterized by a quasi-diaxial orientation of the neighboring hydroxyl groups and fulfill the spatial requirements for hydrogen bonding to the adjacent guanine residues of B-DNA. The striking dependence of potency on the shape of the PAHs is largely caused by repulsion from the C2'-methylene groups of the deoxyribose residues of DNA. This interaction may shift the intercalated triol carbocation, thereby enhancing or reducing the preorganization for covalent bonding. The molecular modeling study is augmented by benchmark ab initio calculations on the bay-region triol carbocation of phenanthrene.

Characterization of polycyclic aromatic hydrocarbons degradative soil Pseudomonas.

Nine Pseudomonas strains, able to degrade polycycle aromatic hydrocarbons (PAHs), were isolated from enriched cultures with naphthalene, as carbon source, and soil samples from a land farming process applied on oil sludge, as inocula. Degradative tests showed that all the strains were capable to catabolize naphthalene (Nah) and phenanthrene (Phn). U2 strain transferred the selected function (Nah) to P. aeruginosa T1 (Hgr Oct+), however some of the transconjugants lost the Oct character, suggesting that it is of plasmidic nature. T1 derivatives as well the wild strains U28 and U31 transferred Nah function to P. putida AC165. All of the examined transconjugants also catabolized phenanthrene, suggesting that Nah and Phn functions in U2, U28, and U31 strains are linked and probably encoded by transferable plasmids.