Aerobic biodegradation of chiral phenoxyalkanoic acid derivatives during incubations with activated sludge

The aerobic biodegradation of racemic mixtures of five chiral phenoxyalkanoic acids was studied according to a biodegradation test that was complemented with enantiomer-specific analysis. Both enantiomers of (RS)-2-phenoxypropanoic acid, (RS)-2-(3-chlorophenoxy)propanoic acid, and (RS)-2-(4-chlorophenoxy)propanoic acid, were completely degraded within 25 days when aerobically incubated with activated sludge. During incubations of (RS)-2-phenoxypropanoic acid, the (R) enantiomer was degraded before the (S) enantiomer, whereas during incubations of (RS)-2-(3-chlorophenoxy)propanoic acid the (S) enantiomer was preferentially degraded. The (R) enantiomer of (RS)-2-(2-chlorophenoxy)propanoic acid was removed after 24 days while only 30% of the (S) enantiomer was degraded within 47 days of incubation. (RS)-2-(2,4,5-Trichlorophenoxy)propanoic acid was the most persistent of all the racemic mixtures tested. After 47 days, the concentration of the (S) enantiomer was nearly unchanged, and the concentration of (R)-2-(2,4,5-trichlorophenoxy)propanoic acid had decreased only by about 40%. The differences observed in the length of the lag phases and in the degradation rates of individual enantiomers can lead to accumulations of the more recalcitrant enantiomer in aquatic or terrestrial ecosystem

ipso-Substitution – A Novel Pathway for Microbial Metabolism of Endocrine-Disrupting 4-Nonylphenols, 4-Alkoxyphenols, and Bisphenol A

Our studies with Sphingobium xenophagum Bayram show that this bacterial strain degrades ?-quaternary 4-nonylphenols by an ipso-substitution mechanism, whereby the nonylphenol substrates are initially hydroxylated at the ipso position to form 4-hydroxy-4-nonylcyclohexa-2,5-dienones (quinols). Subsequently, the ?-quaternary side chains are able to detach as short-living cations from these intermediates. Alkyl branches attached to the carbocation help to delocalize and thereby stabilize the positive charge through inductive and hyperconjugative effects, which explains why only alkyl moieties of ?-quaternary nonylphenols are released. This view is corroborated by experiments with S. xenophagum Bayram, in which the alkyl chains of the non-?-quaternary 4-(1-methyloctyl) phenol (4-NP2) and 4-n-nonylphenol (4-NP1) were not released, so that the bacterium was unable to utilize these isomers as growth substrates. Analysis of dead end metabolites and experiments with 18O labeled H2O and O2 clearly show that in the main degradation pathway the nonyl cation derived from ?-quaternary quinols preferentially combines with a molecule of water to yield the corresponding alcohol and hydroquinone. However, the incorporation of significant amounts of O2-derived oxygen into the nonanol metabolites derived from degradation of certain ?,?-dimethyl substituted nonylphenols by strain Bayram strongly indicates the existence of a minor pathway in which the cation undergoes an alternative reaction and attacks the ipso-hydroxy group, yielding a 4-alkoxyphenol as an intermediate. Additional growth experiments with strain Bayram revealed that also the two alkoxyphenols 4-tert-butoxyphenol and 4-n-octyloxyphenol promote growth. Furthermore, strain Bayram's ipso-hydroxlating activity is able to transform also bisphenol A

Environmental Fate of Chiral Pollutants – the Necessity of Considering Stereochemistry

Many organic compounds regulated by environmental laws are chiral and are released into the environment as racemates. 3-Phenylbutanoic acid and mecoprop, two chiral pollutants, were enantioselectively degraded by pure cultures of microorganisms. This indicates the importance of assessing the environmental impact of stereoisomers separately, because selective enrichment of one of the enantiomers may occur in the environment. Field studies on the fate of highly polar, chiral compounds, like sulfophenylcarboxylates, are hampered by the lack of appropriate analytical methods for the separation of the enantiomers. Therefore, a method based on capillary electrophoresis with α-cyclodextrin as chiral selector was developed to separate the enantiomers of such compounds. In a field study at a Swiss waste disposal site, the fate of the chiral herbicide mecoprop was investigated. The enantiomeric ratio of (R)-mecoprop to (S)-mecoprop altered during groundwater passage of landfill leachate. This is a strong indication for in situ biodegradation. Our data imply that not only the enantiomers of a chiral drug or pesticide may exert different effects on the biological targets, but also their biodegradation and environmental fate may differ

Purification and characterization of hydroquinone dioxygenase from Sphingomonas sp. strain TTNP3

Hydroquinone-1,2-dioxygenase, an enzyme involved in the degradation of alkylphenols in Sphingomonas sp. strain TTNP3 was purified to apparent homogeneity. The extradiol dioxygenase catalyzed the ring fission of hydroquinone to 4-hydroxymuconic semialdehyde and the degradation of chlorinated and several alkylated hydroquinones. The activity of 1 mg of the purified enzyme with unsubstituted hydroquinone was 6.1 μmol per minute, the apparent Km 2.2 μM. ICP-MS analysis revealed an iron content of 1.4 moles per mole enzyme. The enzyme lost activity upon exposure to oxygen, but could be reactivated by Fe(II) in presence of ascorbate. SDS-PAGE analysis of the purified enzyme yielded two bands of an apparent size of 38 kDa and 19 kDa, respectively. Data from MALDI-TOF analyses of peptides of the respective bands matched with the deduced amino acid sequences of two neighboring open reading frames found in genomic DNA of Sphingomonas sp strain TTNP3. The deduced amino acid sequences showed 62% and 47% identity to the large and small subunit of hydroquinone dioxygenase from Pseudomonas fluorescens strain ACB, respectively. This heterotetrameric enzyme is the first of its kind found in a strain of the genus Sphingomonas sensu latu