laccase and 1-aminobenzotriazole

The white rot basidiomycete Ganoderma lucidum was evaluated for its capability to tolerate and to degrade the herbicide diuron. Diuron at a subtoxic concentration was added at the start of the cultivation in glucose liquid stationary cultures. Under this condition diuron was a laccase inducer. Almost 50% of the initially present diuron was removed after 15 d of cultivation. Two diuron metabolites were found N'-(3,4-dichlorophenyl)-N-methylurea (DCPMU) and 3,4-dichlorophenylurea (DCPU). The addition of the cytochrome P450 inhibitors 1-aminobenzotriazole and piperonyl butoxide reduced significantly the capability of the fungus in degrading diuron. The activities of superoxide dismutase and catalase were significantly increased in the mycelial extracts by the presence of diuron. On the other hand, diuron did not cause any significant alteration in the levels of reactive oxygen species. Additionally, laccase could also degrade diuron in vitro and this degradation was increased by the addition of synthetic mediators, 3-ethylbenzthiazoline-6-sulphonic acid and acetylacetone. Significant reduction in the toxicity, as evaluated by the Lactuca sativa bioassay, was observed after G. lucidum treatment. In conclusion, G. lucidum is able to metabolize diuron by intra- and extracellular mechanisms, without the accumulation of toxic products.

Topics: Biotransformation; Catalase; Diuron; Drug Resistance, Fungal; Herbicides; Laccase; Pentanones; Piperonyl Butoxide; Reactive Oxygen Species; Reishi; Superoxide Dismutase; Triazoles

This study examined the interrelation between the biodegradation of polychlorinated biphenyls (PCBs) by Ceriporia sp. ZLY-2010 and its fungal enzyme systems. The degradation rates of Aroclor 1254 and 1260 were 29.01% on day 5 and 36.80% on day 10, respectively. MnP (Manganese dependent peroxidase) and laccase activities showed the greatest increases in the samples containing Aroclors, indicating that extracellular enzymes of Ceriporia sp. ZLY-2010 were affected by the addition of Aroclors. However, the relationship between the biodegradation rate and extracellular enzymes might be obscured by the complexity of the biodegradation process. Cytochrome P450 monooxygenase was inhibited and the biodegradation rate of the Aroclor decreased by adding the inhibitor 1-aminobenzotriazole. Two-dimensional gel electrophoresis showed that intracellular enzymes play a significant role in the biodegradation of Aroclor. Complex extracellular and intracellular enzyme systems in Ceriporia sp. ZLY-2010 play an important role in degrading PCBs. Physiological changes of Ceriporia sp. ZLY-2010 caused by PCBs appeared to affect biodegradation of PCBs. However, it is necessary to further study the unidentified enzymes related to the biodegradation of Aroclor.

Topics: Aroclors; Biodegradation, Environmental; Coriolaceae; Cytochrome P-450 Enzyme System; Enzyme Inhibitors; Laccase; Peroxidases; Polychlorinated Biphenyls; Triazoles

More than 90% of the antibiotics ciprofloxacin (CIPRO) and norfloxacin (NOR) at 2 mg L(-1) were degraded by Trametes versicolor after 7 days of incubation in malt extract liquid medium. In in vitro assays with purified laccase (16.7 nkat mL(-1)), an extracellular enzyme excreted constitutively by this fungus, 16% of CIPRO was removed after 20 h. The addition of the laccase mediator 2,2-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt led to 97.7% and 33.7% degradation of CIPRO and NOR, respectively. Inhibition of CIPRO and NOR degradation by the cytochrome P450 inhibitor 1-aminobenzotriazole suggests that the P450 system also plays a role in the degradation of the two antibiotics. Transformation products of CIPRO and NOR were monitored at different incubation times by triple-quadrupole and quadrupole time-of-flight mass spectrometry, and can be assigned to three different reaction pathways: (i) oxidation of the piperazinyl substituent, (ii) monohydroxylation, and (iii) formation of dimeric products.

Topics: Anti-Bacterial Agents; Biodegradation, Environmental; Biotechnology; Biotransformation; Chromatography, Liquid; Ciprofloxacin; Dimerization; Laccase; Mass Spectrometry; Models, Chemical; Norfloxacin; Oxygen; Tandem Mass Spectrometry; Technology, Pharmaceutical; Time Factors; Trametes; Triazoles

The white-rot fungus Trametes vesicolor degraded naproxen (10 mg L(-1)) in a liquid medium to non-detectable levels after 6h. When naproxen was added in the range of concentrations typically found in the environment (55 microg L(-1)), it was almost completely degraded (95%) after 5h. In vitro degradation experiments with purified laccase and purified laccase plus mediator 1-hydroxybenzotriazol showed slight and almost complete naproxen degradation, respectively. A noticeable inhibition on naproxen degradation was also observed when the cytochrome P450 inhibitor 1-aminobenzotriazole was added to the fungal cultures. These data suggest that both enzymatic systems could play a role in naproxen degradation. 2-(6-hydroxynaphthalen-2-yl)propanoic acid and 1-(6-methoxynaphthalen-2-yl)ethanone were structurally elucidated by HPLC-DAD-MS and NMR as degradation intermediates of naproxen. After 6h of incubation, both parent compound and intermediates disappeared from the medium. The non-toxicity of the treated medium was confirmed by Microtox test.

Topics: Aliivibrio fischeri; Analgesics; Biodegradation, Environmental; Chromatography, High Pressure Liquid; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Laccase; Magnetic Resonance Spectroscopy; Mass Spectrometry; Metabolic Networks and Pathways; Naproxen; Time Factors; Toxicity Tests; Trametes; Triazoles

The laccases of Pycnoporus cinnabarinus and Myceliophthora thermophila are extracellular enzymes with high protein stability. They were used for the 'one pot' synthesis of azole derivatives from 1-aminobenzotriazole together with the p-dihydroxylated laccase substrates 2,5-dihydroxybenzoic acid methyl ester and 2,5-dihydroxybenzoic acid ethyl ester. The reactions yielded heteromolecular dimers (in yields of up to 34%). Methanol was used as the co-solvent to determine the influence of solvent concentration on the course of reaction. The resulting products were isolated, structurally characterized and tested for their antibacterial, antifungal and cytotoxic activities. The products showed low antimicrobial activity and low cytotoxicity compared with commercial available standard compounds but these variables exceeded those of the initial reactants used for the synthesis. In addition to the synthesis of heteromolecular dimers, oligomers were formed and structurally characterized by LC/MS (liquid chromatography/MS).

Topics: Bacillus subtilis; Cell Line, Tumor; Cell Proliferation; Gentisates; Humans; Laccase; Methanol; Microbial Sensitivity Tests; Mitosporic Fungi; Pycnoporus; Staphylococcus aureus; Triazoles

The white-rot fungus Trametes versicolor degraded trichloroethylene (TCE), a highly oxidized chloroethene, and produced 2,2,2-trichloroethanol and carbon dioxide as the main products of degradation, based on the results obtained using [13C]-TCE as the substrate. For a range of concentrations of TCE between 2 and 20 mg l(-1), 53% of the theoretical maximum chloride expected from complete degradation of TCE was observed. Laccase was shown to be induced by TCE, but did not appear to play a role in TCE degradation. Cytochrome P-450 appears to be involved in TCE degradation, as evidenced by marked inhibition of degradation of TCE in the presence of 1-aminobenzotriazole, a known inhibitor of cytochrome P-450. Our results suggested that chloral (trichloroacetaldehyde) was an intermediate of the TCE degradation pathway. The results indicate that the TCE degradation pathway in T. versicolor appears to be similar to that previously reported in mammals and is mechanistically quite different from bacterial TCE degradation.

Topics: Carbon Dioxide; Chlorides; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Ethylene Chlorohydrin; Laccase; Polyporales; Triazoles; Trichloroethylene; Water Pollutants, Chemical