Gas chromatographic-mass spectrometric determination of chlorinated cis-1,2-dihydroxycyclohexadienes and chlorocatechols as their boronates

Kirsch, Nils; Stan, Hans‐Jürgen

doi:10.1016/0021-9673(94)00619-9

Cited by 13 publications

(17 citation statements)

References 22 publications

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“…Compounds: 1, biphenyl; 2, biphenyl-2,3-dihydrodiol; 3, 2,3-dihydroxybiphenyl; 4, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid; 5a, 2-hydroxypenta-2,4-dienoic acid; 5b, benzoic acid. ions (M-56) ϩ (loss of C 4 H 8 ) were the only prominent positively charged species, indicating the formation of catechols (22). In accordance with this, the loss of one of the substrate halogens was clearly visible.…”

Section: Resultssupporting

confidence: 68%

“…To obtain butylboronate derivatives, the solvent was removed from 200 l of extract, and the residue was redissolved in 80 l of acetone. Twenty milliliters of a 2-mg/ml solution of n-butylboronic acid in acetone was added, and the solution was incubated at 50°C for 10 min (22). To obtain trimethylsilyl (TMS) derivatives, the solvent was removed from 100 l of extract, the residue was redissolved in 50 l of N,O-bis-trimethylsilyltrifluoroacetamide and trimethylchlorosilane (99:1 [vol/vol]), and the solution was incubated at 70°C for 30 min.…”

Section: Methodsmentioning

confidence: 99%

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Dehalogenation, Denitration, Dehydroxylation, and Angular Attack on Substituted Biphenyls and Related Compounds by a Biphenyl Dioxygenase

2001

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The attack by the bph-encoded biphenyl dioxygenase of Burkholderia sp. strain LB400 on a number of symmetrical ortho-substituted biphenyls or quasi ortho-substituted biphenyl analogues has been investigated. 2,2-Difluoro-, 2,2-dibromo-, 2,2-dinitro-, and 2,2-dihydroxybiphenyl were accepted as substrates. Dioxygenation of all of these compounds showed a strong preference for the semisubstituted pair of vicinal ortho and meta carbons, leading to the formation of 2-substituted 2,3-dihydroxybiphenyls by subsequent elimination of HX (X ‫؍‬ F, Br, NO 2 , or OH). All of these products were further metabolized by 2,3-dihydroxybiphenyl 1,2-dioxygenases of Burkholderia sp. strain LB400 or of Rhodococcus globerulus P6. Dibenzofuran and dibenzodioxin, which may be regarded as analogues of doubly ortho-substituted biphenyls or diphenylethers, respectively, were attacked at the "quasi ortho" carbon (the angular position 4a) and its neighbor. This shows that an aromatic ring-hydroxylating dioxygenase of class IIB is able to attack angular carbons. The catechols formed, 2,3,2-trihydroxybiphenyl and 2,3,2-trihydroxydiphenylether, were further metabolized by 2,3-dihydroxybiphenyl 1,2-dioxygenase. While angular attack by the biphenyl dioxygenase was the main route of dibenzodioxin oxidation, lateral dioxygenation leading to dihydrodiols was the major reaction with dibenzofuran. These results indicate that this enzyme is capable of hydroxylating ortho or angular carbons carrying a variety of substituents which exert electron-withdrawing inductive effects. They also support the view that the conversions of phenols into catechols by ring-hydroxylating dioxygenases, such as the transformation of 2,2-dihydroxybiphenyl into 2,3,2-trihydroxybiphenyl, are the results of di-rather than of monooxygenations. Lateral dioxygenation of dibenzofuran and subsequent dehydrogenation and extradiol dioxygenation by a number of biphenyl-degrading strains yielded intensely colored dead-end products. Thus, dibenzofuran can be a useful chromogenic indicator for the activity of the first three enzymes of biphenyl catabolic pathways.Ring-hydroxylating dioxygenases are key enzymes of the aerobic bacterial catabolism of aromatic compounds. Generally, this family of enzymes has been shown to be quite versatile with respect to the substrates accepted as well as to the type of reactions catalyzed (11, 28). The prototype reaction supported is the addition of two hydroxy groups to vicinal carbons of aromatic rings which, usually via a subsequent dehydrogenation, leads to the formation of catechols (Fig. 1). The hydroxy groups enable and direct fission of the aromatic ring by extraor intradiol dioxygenases which cleave the carbon-carbon bonds either between or adjacent to these substituents (Fig. 1). We previously characterized the dioxygenations of chlorinated biphenyls (Cl-Bs) as catalyzed by two bacterial enzymes, the bph-encoded biphenyl dioxygenases (BphAs) of Burkholderia sp. strain LB400 and of Rhodococcus globerulus P6 (26, 33, 35). In the course of those ...

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Section: Resultssupporting

confidence: 68%

Section: Methodsmentioning

confidence: 99%

Dehalogenation, Denitration, Dehydroxylation, and Angular Attack on Substituted Biphenyls and Related Compounds by a Biphenyl Dioxygenase

2001

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“…Whereas no such products were detected from 23DCT and 35DCT, prominent signals showing the expected molecular ion of m/z 260, 262, and 264 (relative intensities, 100:62:9) were observed from 34DCT, 24DCT, 25DCT, and 26DCT. The products showed the fragmentation pattern typical for n-butylboronated chlorobenzene or methylbenzene dihydrodiols (5,16) . Analysis of the boronated extract gave no indication for the formation of dihydroxylated products.…”

Section: Resultsmentioning

confidence: 98%

Transformation of Chlorinated Benzenes and Toluenes by Ralstonia sp. Strain PS12 tecA (Tetrachlorobenzene Dioxygenase) and tecB (Chlorobenzene Dihydrodiol Dehydrogenase) Gene Products

Pollmann

Beil

Pieper

2001

Appl Environ Microbiol

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The tecB gene, located downstream of tecA and encoding tetrachlorobenzene dioxygenase, in Ralstonia sp. strain PS12 was cloned into Escherichia coli DH5␣ together with the tecA gene. The identity of the tecB gene product as a chlorobenzene dihydrodiol dehydrogenase was verified by transformation into the respective catechols of chlorobenzene, the three isomeric dichlorobenzenes, as well as 1,2,3-and 1,2,4-trichlorobenzenes, all of which are transformed by TecA into the respective dihydrodihydroxy derivatives. Di-and trichlorotoluenes were either subject to TecA-mediated dioxygenation (the major or sole reaction observed for the 1,2,4-substituted 2,4-, 2,5-, and 3,4-dichlorotoluenes), resulting in the formation of the dihydrodihydroxy derivatives, or to monooxygenation of the methyl substituent (the major or sole reaction observed for 2,3-, 2,6-, and 3,5-dichloroand 2,4,5-trichlorotoluenes), resulting in formation of the respective benzyl alcohols. All of the chlorotoluenes subject to dioxygenation by TecA were transformed, without intermediate accumulation of dihydrodihydroxy derivatives, into the respective catechols by TecAB, indicating that dehydrogenation is no bottleneck for chlorobenzene or chlorotoluene degradation. However, only those chlorotoluenes subject to a predominant dioxygenation were growth substrates for PS12, confirming that monooxygenation is an unproductive pathway in PS12.

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“…Extraction of resting-cell supernatants and drying and evaporation of solvents were carried out as previously described (33). To obtain butylboronate derivatives, dried extracts were dissolved in 80 l of acetone, 20 l of a 2-mg/ml solution of n-butylboronic acid in acetone was added, and the solution incubated at 50ЊC for 10 min (25). After derivatization, mixtures were evaporated to dryness under a stream of nitrogen and dissolved in 10 or 20 l of n-octane.…”

Section: Methodsmentioning

confidence: 99%

Heterologous expression of biphenyl dioxygenase-encoding genes from a gram-positive broad-spectrum polychlorinated biphenyl degrader and characterization of chlorobiphenyl oxidation by the gene products

et al. 1997

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The bphA1A2A3A4 gene cluster, encoding a biphenyl dioxygenase from Rhodococcus globerulus P6, a grampositive microorganism able to degrade a wide spectrum of polychlorobiphenyls (PCBs), was expressed in Pseudomonas putida, thereby allowing characterization of chlorobiphenyl oxidation by this enzyme. While P6 biphenyl dioxygenase activity was observed in P. putida containing bphA1A2A3A4, no activity was detected in Escherichia coli cells containing the same gene cluster. In E. coli, transcription of genes bphB and bphC1, located downstream of bphA1A2A3A4, was shown to be driven solely by a vector promoter, which indicated that the lack of biphenyl dioxygenase activity was not due to a lack of mRNA synthesis. Radioactive labelling of bph gene products in E. coli implied inefficient translation of the bphA gene cluster or rapid degradation of the gene products. The biosynthesis of functional P6 biphenyl dioxygenase in P. putida cells containing the same plasmid construct that yielded no activity in E. coli emphasizes the importance of the host strain for heterologous expression and shows that synthesis, correct folding, and assembly of a Rhodococcus biphenyl dioxygenase can be achieved in a gram-negative organism. Dioxygenation of six mono-and dichlorinated PCB congeners by P. putida containing the P6 bphA gene cluster indicates the following ring substitution preference for this reaction (from most to least preferred): un-, meta-, para-, and ortho-substitution. No indications were found for dioxygenation of meta/para carbon pairs, or for hydroxylation of chlorinated carbons at any position of a monochlorinated ring, suggesting a strict specificity of this biphenyl dioxygenase for attack at nonhalogenated ortho/meta vicinal carbons. This contrasts the properties of an analogous enzyme from Pseudomonas sp. strain LB400, which can both dioxygenate at meta and para positions and dehalogenate substituted ortho carbons during ortho and meta dioxygenation.

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Gas chromatographic-mass spectrometric determination of chlorinated cis-1,2-dihydroxycyclohexadienes and chlorocatechols as their boronates

Cited by 13 publications

References 22 publications

Dehalogenation, Denitration, Dehydroxylation, and Angular Attack on Substituted Biphenyls and Related Compounds by a Biphenyl Dioxygenase

Dehalogenation, Denitration, Dehydroxylation, and Angular Attack on Substituted Biphenyls and Related Compounds by a Biphenyl Dioxygenase

Transformation of Chlorinated Benzenes and Toluenes by Ralstonia sp. Strain PS12 tecA (Tetrachlorobenzene Dioxygenase) and tecB (Chlorobenzene Dihydrodiol Dehydrogenase) Gene Products

Heterologous expression of biphenyl dioxygenase-encoding genes from a gram-positive broad-spectrum polychlorinated biphenyl degrader and characterization of chlorobiphenyl oxidation by the gene products

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