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Halophenols and their derivatives are priority pollutants of mainly anthropogenic origin. Over several decades, these compounds have been widely used as building blocks in chemical and pharmaceutical syntheses and as herbicides and pesticides, and they have caused serious local contamination of the environment. Soil microorganisms have developed the capacity of utilizing halophenols for their growth by a diverse set of biodegradation pathways (8). Aerobic soil microorganisms generally degrade mono-and dihalophenols through the initial action of (chloro)phenol ortho-hydroxylases, leading to the formation of halocatechols (1,7,9,10,12). In the framework of a project devoted to the biodegradation of halophenols by gram-positive bacteria, we investigated the formation of hydroxylated intermediates formed upon the conversion of halophenols by various Rhodococcus species and previously demonstrated the formation of (halo)catechols as initial intermediates in the biodegradation pathways (3). However, identification of the subsequent biodegradation pathways of the chlorocatechols appeared hampered by the fact that unequivocal identification of the site of introduction of a third hydroxyl group is difficult because 1 H nuclear magnetic resonance (NMR) splitting patterns combined with 1 H chemical shift data of the protons present in these metabolites can be compatible with more than one substitution pattern (13). Therefore, in this paper, we have studied the possible formation of trihydroxyfluorobenzene metabolites from fluorophenols by whole cells of Rhodococcus opacus 1cp in detail. The fluorine substituent provides the possibility to detect and quantify the possible hydroxyfluorobenzene intermediates by 19 F NMR, allowing the identification of the exact substitution pattern. Using this technique we unambiguously demonstrate the formation of fluoropyrogallols (1,2,3-trihydroxyfluorobenzenes) as new intermediates in the biotransformation of monofluorophenols by R. opacus 1cp. MATERIALS AND METHODS Chemicals.Phenol was purchased from Merck (Darmstadt, Germany). 2-Fluorophenol, 3-fluorophenol, and 4-fluorophenol were purchased from Janssen Chimica (Beerse, Belgium). Fluorocatechols were prepared from the corresponding fluorophenols using purified phenol hydroxylase from Trichosporon cutaneum (14). Fluoromuconates were prepared and identified as described previously (2) by incubating the fluorocatechols with catechol 1,2-dioxygenase from Pseudomonas arvilla C-1.Growth of R. opacus 1cp. The strain R. opacus 1cp was isolated and maintained as described previously (6). The strain can grow on phenol as the sole source of carbon. For cultivation, a mineral synthetic medium containing, per liter, 1 g of NH 4 NO 3 , 1 g of K 2 HPO 4 , 1 g of KH 2 PO 4 , 0.2 g of MgSO 4 ⅐ 7H 2 O, 0.02 g of CaCl 2 , and 2 drops of a saturated solution of FeCl 3 (pH 7.2) was used. Phenol was used as the source of carbon and was initially added at a 200-mg/liter final concentration. R. opacus 1cp did not grow on either of the three monofluorophenols ...

Section: Conversion Of Monofluorophenols By R Opacus 1cpsupporting

confidence: 65%

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confidence: 99%

Identification of Fluoropyrogallols as New Intermediates in Biotransformation of Monofluorophenols in Rhodococcus opacus 1cp

Finkelstein

Баскунов

Boersma

et al. 2000

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“…Examples of biodegradation of fluorinated compounds most commonly found in the literature involve fluorobenzoic acids (7,15,20,21,24) and fluorophenols (1,2,23). Although degradation under aerobic conditions is usually reported, anaerobic degradation of fluorobenzoates under denitrifying conditions has also been reported (26).…”

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confidence: 99%

“…Studies on the metabolism of 2-flurobenzoate have shown that cleavage of the C-F bond occurs incidentally during oxygenase attack on the aromatic ring (20). It has been reported that biodegradation of fluorophenol occurs through a phenol hydroxylase (2). Some authors suggested that 3-fluorobenzoate, 4-fluorobenzoate, and difluorobenzoate are metabolized via 4-fluorocatechol (4,5,24).…”

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confidence: 99%

Isolation and Initial Characterization of a Bacterial Consortium Able To Mineralize Fluorobenzene

Carvalho

Alves

Ferreira

et al. 2002

Fluorinated compounds are known to be more resistant to microbial degradation than other halogenated chemicals. A microbial consortium capable of aerobic biodegradation of fluorobenzene (FB) as the sole source of carbon and energy was isolated by selective enrichment from sediments collected in a drain near an industrial site. A combination of three microbial strains recovered from the enriched consortium was shown to be necessary for complete FB mineralization. Two of the strains (F1 and F3) were classified by 16S rRNA analysis as belonging to the Sphingobacterium/Flavobacterium group, while the third (F4) falls in the ␤-Proteobacteria group, clustering with Alcaligenes species. Strain F4 was consistently found in the liquid cultures in a much greater proportion than strains F1 and F3 (86:8:6 for F4, F1, and F3, respectively). Stoichiometric release of fluoride ions was measured in batch and fed-batch cultures. In batch cultures, the consortium was able to use FB up to concentrations of 400 mg liter ؊1 and was able to utilize a range of other organic compounds, including 4-fluorophenol and 4-fluorobenzoate. To our knowledge this is the first time biodegradation of FB as a sole carbon source has been reported.The advances in organic synthesis have led to the introduction of numerous new organic compounds into the environment, whose susceptibilities to biotreatment processes are unknown. Fluoroaromatics are being increasingly used in a wide range of agrochemical and pharmaceutical products, due to the need to find environmentally acceptable alternatives to chlorinated compounds (17). The diversity of structures and the chemical inertness of many halogenated organics pose particular problems and challenges for microbial degradation (10). Some authors propose that the recalcitrance of a halogenated organic compound usually becomes greater with the increase of the electronegativity of the substituents; thus, the recalcitrance of F-C is greater than that of Cl-C, Br-C, and I-C (9).The biodegradation of a vast range of halogenated aromatic compounds, especially chlorinated compounds, has been described (13,22), but scant information is available on the metabolic and cometabolic fate of fluorinated aromatic compounds in bacteria. Examples of biodegradation of fluorinated compounds most commonly found in the literature involve fluorobenzoic acids (7,15,20,21,24) and fluorophenols (1, 2, 23). Although degradation under aerobic conditions is usually reported, anaerobic degradation of fluorobenzoates under denitrifying conditions has also been reported (26). The existence of various metabolic pathways, some of which may lead to the formation of inhibitor metabolites, has been reported (15,24,25). In some cases, as in the degradation of fluoroacetate, a specific enzyme is responsible for the cleavage of the C-F bond (12). Studies on the metabolism of 2-flurobenzoate have shown that cleavage of the C-F bond occurs incidentally during oxygenase attack on the aromatic ring (20). It has been reported that biodegradation of fluo...

“…To that end, several fluoromuconates were enzymatically produced from halophenols and 19 F nuclear magnetic resonance (NMR) was used for the identification of reaction products. The 19 F NMR technique combines high selectivity with high sensitivity and is especially useful for the in situ detection of unstable intermediates (2,4,5,6,24). It is shown for the first time that 2-fluoromuconate is converted by ClcB2 to 5-fluoromuconolactone and that the latter compound is dehalogenated by ClcF to cis-dienelactone.…”

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confidence: 99%

Conversion of 2-Fluoromuconate tocis-Dienelactone by Purified Enzymes ofRhodococcus opacus1cp

Solyanikova

Moiseeva

Boeren³

et al. 2003

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The present study describes the 19 F nuclear magnetic resonance analysis of the conversion of 3-halocatechols to lactones by purified chlorocatechol 1,2-dioxygenase (ClcA2), chloromuconate cycloisomerase (ClcB2), and chloromuconolactone dehalogenase (ClcF) from Rhodococcus opacus 1cp grown on 2-chlorophenol. The 3-halocatechol substrates were produced from the corresponding 2-halophenols by either phenol hydroxylase from Trichosporon cutaneum or 2-hydroxybiphenyl 3-mono-oxygenase from Pseudomonas azelaica.