Breakdown products on metabolic pathway of degradation of benz[a]anthracene by a ligninolytic fungus

Cultures of the ligninolytic fungus Irpex lacteus incubated in a nutrient liquid medium degraded more than 70% of the initially applied benz[a]anthracene within 14 days. At the first step of metabolization, benz[a]anthracene was transformed via a typical pathway of ligninolytic fungi to benz[a]anthracene-7,12-dione (BaAQ). The product was further transformed by at least two ways, whereas one is complied with the anthracene metabolic pathway of I. lacteus. Benz[a]anthracene-7,12-dione was degraded to 1,2-naphthalenedicarboxylic acid and phthalic acid that was followed with production of 2-hydroxymethyl benzoic acid or monomethyl and dimethylesters of phthalic acid. Another degradation product of BaAQ was identified as 1-tetralone. Its transformation via 1,4-naphthalenedione, 1,4-naphthalenediol and 1,2,3,4-tetrahydro-1-hydroxynaphthalene resulted again in phthalic acid. None of the intermediates were identified as dead-end metabolites. Metabolites produced by ring cleavage of benz[a]anthracene using the ligninolytic fungus are firstly presented in this work.     

Phenanthrene degradation in Arthrobacter sp. P1-1: initial 1,2-, 3,4- and 9,10-dioxygenation, and meta- and ortho-cleavages of naphthalene-1,2-diol after its formation from naphthalene-1,2-dicarboxylic acid and hydroxyl naphthoic acids

Arthrobacter sp. P1-1, isolated from a polycyclic aromatic hydrocarbon (PAH)-contaminated site in Hilo, HI, USA, can decompose phenanthrene (40 mg l⁻¹) completely within 7 days. A detailed phenanthrene metabolism map was constructed based on metabolite analysis and replacement cultures. Initial dioxygenation occurs on 1,2-, 3,4-, and 9,10-C of phenanthrene, dominantly on 3,4-C positions. Rapid accumulation of 5,6- and 7,8-benzocoumarin suggests that phenanthrene-1,2- and -3,4-diols mainly undergo meta-cleavage. However, a trace amount of o-carboxyvinylnaphthoates and diphenic acid indicates a limited extent of ortho-cleavage of the diols. Naphthalene-1,2-diol, as a common and converged metabolite, was formed from 1-[(E)-2-carboxyvinyl]-2-naphthoic acid, naphthalene-1,2-dicarboxylic acid, and 1-hydroxy-2-naphthoic acid in separate culture tests. Naphthalene-1,2-diol is then degraded in a dominant phthalic acid pathway and a minor salicylic acid pathway. Several metabolites of phthalic acid were found, while no salicylic acid metabolites were detected. The strain P1-1 likely has a very diverse set of PAH-degrading enzymes or the enzymes having relaxed substrate-specificity.     

Complete degradation of butyl benzyl phthalate by a defined bacterial consortium: role of individual isolates in the assimilation pathway

Two bacterial strains, in consortium, were isolated by enrichment techniques from municipal waste-contaminated soil, which utilized butyl benzyl phthalate (BBP) as the sole carbon source. One of the isolates was identified as Arthrobacter sp. strain WY and the other one as Acinetobacter sp. strain FW based on the morphological, nutritional and biochemical characteristics and 16S rRNA sequence analysis. Various metabolites of BBP engendered by Arthrobacter sp. strain WY were isolated and identified by a combination of chromatographic and spectrophotometric analyses, which revealed a pathway involving monobutylphthalate (MBuP), monobenzyl phthalate (MBzP), phthalic acid and protocatechuic acid. The protocatechuic acid in turn was processed by ortho-cleavage dioxygenase to form beta-carboxy-cis,cis-muconate, ultimately leading to the TCA cycle. The Arthrobacter sp. strain WY could not utilize the hydrolyzed alcohols of BBP. On the other hand, the Acinetobacter sp. strain FW, which by itself could not utilize BBP as the sole carbon source, is capable of utilizing the hydrolyzed alcohols of BBP. Benzyl alcohol was found to be metabolized by the Acinetobacter sp. strain FW via benzaldehyde, benzoic acid and catechol. Catechol was further degraded by ortho-cleavage dioxygenase to cis,cis-muconic acid and subsequently to muconolactone leading to beta-ketoadipate pathway. Moreover, the Acinetobacter sp. strain FW metabolized 1-butanol through butyraldehyde and butyric acid leading to the tricarboxylic acid cycle via beta-oxidation pathway. This is the first report on the complete degradation of BBP by a defined consortium describing the role of its individual constituents in the BBP assimilation pathway.     

Study of metabolites from the degradation of polycyclic aromatic hydrocarbons (PAHs) by bacterial consortium enriched from mangrove sediments

The PAH metabolites produced during degradation of fluorene, phenanthrene and pyrene by a bacterial consortium enriched from mangrove sediments were analyzed using the on-fiber silylation solid-phase microextraction (SPME) combining with gas chromatography–mass spectrometry (GC–MS) method. Seventeen metabolites at trace levels were identified in different PAH degradation cultures based on the full scan mass spectra. In fluorene degradation cultures, 1-, 2-, 3- and 9-hydroxyfluorene, fluorenone, and phthalic acid were detected. In phenanthrene and pyrene degradation cultures, various common metabolites such as phenanthrene and pyrene dihydrodiols, mono-hydroxy phenanthrene, dihydroxy pyrene, lactone and 4-hydroxyphenanthrene, methyl ester, and phthalic acid were found. The detection of various common and novel metabolites demonstrates that SPME combining with GC–MS is a quick and convenient method for identification as well as monitoring the real time changes of metabolite concentrations throughout the degradation processes. The knowledge of PAH metabolic pathways and kinetics within indigenous bacterial consortium enriched from mangrove sediments contributes to enhance the bioremediation efficiency of PAH in real environment.     

Novel degradation pathway of 4-chloro-2-aminophenol via 4-chlorocatechol in Burkholderia sp. RKJ 800

Burkholderia sp. RKJ 800 utilized 4-chloro-2-aminophenol (4C2AP) as the sole carbon and energy source and degraded it with release of chloride and ammonium ions. The metabolic pathway of degradation of 4C2AP was studied and a novel intermediate, 4-chlorocatechol was identified as a major degradation product of 4C2AP using high-performance liquid chromatography and gas chromatography–mass spectrometry. Enzyme activities for 4C2AP-deaminase and 4-chlorocatechol-1,2-dioxygenase were detected in the crude extracts of the 4C2AP-induced cells of strain RKJ 800. The activity of the 4C2AP-deaminase confirmed the formation of 4-chlorocatechol from 4C2AP and the 4-chlorocatechol-1,2-dioxygenase activity suggested the cleavage of 4-chlorocatechol into 3-chloro-cis,cis-muconate. On the basis of the identified metabolites, we have proposed a novel degradation pathway of 4C2AP for Burkholderia sp. RKJ 800. Furthermore, the potential of Burkholderia sp. RKJ 800 to degrade 4C2AP in soil was also investigated using microcosm studies under laboratory conditions. The results of microcosm studies conclude that Burkholderia sp. RKJ 800 was able to degrade 4C2AP in soil and may be used to remediate 4C2AP-contaminated site. This is the first report of (1) the formation of 4-chlorocatechol and 3-chloro-cis,cis-muconate in the degradation pathway of 4C2AP and (2) bioremediation of 4C2AP by any bacterium.     

CO(2)-H(2)-dependent anaerobic biotransformation of phthalic acid isomers in sediment slurries

This paper investigates the anaerobic biotransformation of three isomers of phthalic acid and benzoic acid in sediment slurries under four different atmospheres [N(2), N(2)/H(2) (19:1, v/v), CO(2), and CO(2)/H(2) (4:1, v/v)]. Significant differences were observed in lag periods and biotransformation rates among the phthalic acid isomers and under the different atmospheres. In most cases, the relative biotransformation rates of the three isomers of phthalic acid were ortho-phthalic acid>isophthalic acid>terephthalic acid. Benzoate was transformed faster than any isomer of phthalic acid. Since biotransformation of phthalic acid isomers in sediment slurries was enhanced by high initial levels of H(2) and CO(2) in the headspace, we propose a pathway for phthalic acid biodegradation in which the initial transformation to benzoate is CO(2)-H(2) dependent. Acetogenic bacteria were investigated for their possible involvement in this transformation reaction, but when MPN counts were used to compare the growth dynamics of acetogenic bacteria with the time course of the terephthalic acid transformation under N(2)/H(2) (19:1, v/v) and CO(2)/H(2) (4:1, v/v) atmospheres, the results were inconclusive.     

Complete degradation of di-n-octyl phthalate by Gordonia sp. strain Dop5

The present study describes the assimilation of di-n-octyl phthalate by an aerobic bacterium, isolated from municipal waste-contaminated soil sample utilizing di-n-octyl phthalate as the sole source of carbon and energy. The isolate was identified as Gordonia sp. based on the morphological, nutritional and biochemical characteristics as well as 16S rRNA gene sequence analysis. A combination of chromatographic and spectrometric analyses revealed a complete di-n-octyl assimilation pathway. In the degradation process, mono-n-octyl phthalate, phthalic acid, protocatechuic acid and 1-octanol were identified as the degradation products of di-n-octyl phthalate. Furthermore, phthalic acid was metabolized via protocatechuic acid involving protocatechuate 3,4-dioxygenase while 1-octanol was metabolized by NAD⁺-dependent dehydrogenases to 1-octanoic acid, which was subsequently degraded via β-oxidation, ultimately, leading to tricarboxylic acid cycle intermediates. Apart from phthalic acid and 1-octanol metabolizing pathway enzymes, two esterases, di-n-octyl phthalate hydrolase and mono-n-octyl phthalate hydrolase involved in di-n-octyl phthalate degradation were found to be inducible in nature. This is the first report on the metabolic pathway involved in the complete degradation of di-n-octyl phthalate by a single bacterial isolate, which is also capable of efficiently degrading other phthalate esters of environmental concern having either shorter or longer alkyl chains.     

Degradation of 3,4-dichloro- and 3,4-difluoroaniline by Pseudomonas fluorescens 26-K

3,4-Dichloro- and 3,4-difluoroanilines were degraded by Pseudomonas fluorescens 26-K under aerobic conditions. In the presence of glucose strain degraded 170 mg/L of 3,4-dichloroaniline (3,4-DCA) during 2-3 days. Increasing of toxicant concentration up to 250 mg/L led to degradation of 3,4-DCA during 4 days and its intermediates during 5-7 days. Without cosubstrate and nitrogen source degradation of 3,4-DCA took place too, but more slowly--about 40% of toxicant at initial concentration 75 mg/L was degraded during 15 days. 3,4-Difluoroaniline (3,4-DFA) (initial concentration 170 mg/L) was degraded by Pseudomonas fluorescens 26-K during 5-7 days. The strain was able to completely degrade up to 90 mg/L of 3,4-DFA, without addition of cosubstrate and nitrogen during 15 days. Degradation of fluorinated aniline was accompanied by intensive defluorination. Activity of catechol 2,3-dioxygenase (C2,3DO) (0.230 micromol/min/mg of protein) was found in the culture liquid of the strain, grown with 3,4-DCA and glucose. This fact, as well as, the presence of 3-chloro-4-hydroxyaniline as a metabolite suggested that 3,4-DCA degradation pathway includes dehalogenation and hydroxylation of aromatic ring followed by its subsequent cleaving by C2,3DO. On the contrary, activity of catechol 1,2-dioxygenase (C1,2DO) (0.08 micromol/min/mg of protein) was found in the cell-free extract of biomass grown on 3,4-DFA. 3-Fluoro-4-hydroxyaniline as intermediate was found in this cell-free extract.     

Sonochemical substrate selectivity and reaction pathway of systematically substituted azo compounds

The sonochemical degradation of the systematically substituted azo compound 2,7-dihydroxy-1-phenylazonaphthaline-3,6-disulfonic acid was investigated using a frequency of 850 kHz and an acoustic input power of 61 W. All derivatives were degraded completely within 6h by the ultrasonic treatment. Trifluoromethyl substituted azo compounds exhibited 2-3-fold higher degradation rates in comparison to the reference hydrogen substituted azo compound (k=0.54 h(-1)). In contrast to enzymatic processes (azoreductase or laccase), the ultrasonic treatment for these ortho-, meta-, and para-substituted azo compound showed 1.5-50-fold higher degradation rates. Additionally the ultrasound treatment was characterized by shorter reaction times. As a result of the detection and identification of specific intermediates using LC-MS a reaction pathway of the sonochemical degradation of the analysed azo compound is proposed indicating the formation of cyclohexadienone and naphthalene quinone derivatives.     

一株1，2-二氯苯降解菌的分离鉴定及其降解特性

采用富集驯化方法,从盐城芦苇湿地根际土壤中分离得到一株可高效降解1,2-二氯苯的菌株,命名为DL-1。该菌株可以在以1,2-二氯苯为惟一碳源的无机培养基上生长,能够耐受最高浓度为200 mg/L的1,2-二氯苯。根据形态特征观察、生理生化鉴定和16S rDNA序列同源性分析,该目标菌株被鉴定为蜡质芽孢杆菌（Bacillus cereus）。菌株DL-1对1,2-二氯苯降解性能研究表明,该菌株为一株兼性厌氧菌,其适宜降解浓度、适宜温度、适宜pH值和适宜接种量分别为120mg/L、32℃、7和10%,在适宜降解条件下降解12,-二氯苯4 d其降解率达到80.3%。本实验为利用该菌株降解12,-二氯苯污水的应用提供了理论基础。     

Biodegradation of nonylphenol in river sediment

We investigated the biodegradation of nonylphenol monoethoxylate (NP1EO) and nonylphenol (NP) by aerobic microbes in sediment samples collected at four sites along the Erren River in southern Taiwan. Aerobic degradation rate constants (k1) and half-lives (t1/2) for NP (2 microg g(-1)) ranged from 0.007 to 0.051 day(-1) and 13.6 to 99.0 days, respectively; for NP1EO (2 microg g(-1)) the ranges were 0.006 to 0.010 day(-1) and 69.3 to 115.5 days. Aerobic degradation rates for NP and NP1EO were enhanced by shaking and increased temperature, and delayed by the addition of Pb, Cd, Cu, Zn, phthalic acid esters (PAEs), and NaCl, as well as by reduced levels of ammonium, phosphate, and sulfate. Of the microorganism strains isolated from the sediment samples, we found that strain JC1 (identified as Pseudomonas sp.) expressed the best biodegrading ability. Also noted was the presence of 4'-amino-acetophenone, an intermediate product resulting from the aerobic degradation of NP by Pseudomonas sp.     

Mechanisms of biodegradation of dibenzoate plasticizers

Biodegradation mechanisms were elucidated for three dibenzoate plasticizers: diethylene glycol dibenzoate (D(EG)DB), dipropylene glycol dibenzoate (D(PG)DB), both of which are commercially available, and 1,6-hexanediol dibenzoate, a potential green plasticizer. Degradation studies were done using Rhodococcus rhodochrous in the presence of pure alkanes as a co-substrate. As expected, the first degradation step for all of these systems was the hydrolysis of one ester bond with the release of benzoic acid and a monoester. Subsequent biodegradation of the monobenzoates of diethylene glycol (D(EG)MB) and dipropylene glycol (D(PG)MB) was very slow, leading to significant accumulation of these monoesters. In contrast, 1,6-hexanediol monobenzoate was quickly degraded and characterization of the metabolites indicated that the biodegradation proceeded by way of the oxidation of the alcohol group to generate 6-(benzoyloxy) hexanoic acid followed by β-oxidation steps. This pathway was blocked for D(EG)MB and D(PG)MB by the presence of an ether function.The use of a pure hydrocarbon as a co-substrate resulted in the formation of another class of metabolites; namely the esters of the alcohols formed by the oxidation of the alkanes and the benzoic acid released by hydrolysis of the original diesters. These metabolites were biodegraded without the accumulation of any intermediates.     

Biodegradation of aromatic hydrocarbons by Haloarchaea and their use for the reduction of the chemical oxygen demand of hypersaline petroleum produced water

Ten halophilic Archaea (Haloarchaea) strains able to degrade aromatic compounds were isolated from five hypersaline locations; salt marshes in the Uyuni salt flats in Bolivia, crystallizer ponds in Chile and Cabo Rojo (Puerto Rico), and sabkhas (salt flats) in the Persian Gulf (Saudi Arabia) and the Dead Sea (Israel and Jordan). Phylogenetic identification of the isolates was determined by 16S rRNA gene sequence analysis. The isolated Haloarchaea strains were able to grow on a mixture of benzoic acid, p-hydroxybenzoic acid, and salicylic acid (1.5 mM each) and a mixture of the polycyclic aromatic hydrocarbons, naphthalene, anthracene, phenanthrene, pyrene and benzo[a]anthracene (0.3 mM each). Evaluation of the extent of degradation of the mixed aromatic hydrocarbons demonstrated that the isolates could degrade these compounds in hypersaline media containing 20% NaCl. The strains were shown to reduce the COD of hypersaline crude oil reservoir produced waters significantly beyond that achieved using standard hydrogen peroxide treatment alone.     

Phylogenetic and degradation characterization of Burkholderia cepacia WZ1 degrading herbicide quinclorac

Strain WZI capable of degrading quinclorac was isolated from a pesticide manufactory soil and considered to be Burkholderia cepacia, belonged to bacteria, Proteobacteria, beta-Proteobacteria, based on morphology, physio-biochemical properties, whole cell fatty acid analysis and a partial sequencing of 16S rDNA. Strain WZ1 decomposed 90% of quinclorac at original concentration of 1000 mg L(-1) within 11 days. GC/MS analysis showed that the strain degraded quinclorac to 3,7-dichloro-8-quinoline and the cracked residue 2-chloro, 1,4-benzenedicarboxylic acid, indicating that the metabolic pathway was initiated by process of decarboxylation followed by cleavage of the aromatic ring. Stain WZ1 was also able to degrade some other herbicides and aromatic compounds, including 2,4,5-T, phenol, naphthalene and hydrochinone etc. This paper describes for the first time Phylogenetic and degradation characterization of a pure bacterium which, is able to mineralize quinclorac.     

铜绿假单胞菌GF31降解土壤中氯氰菊酯的性能研究

利用实验室分离获得铜绿假单胞菌GF31(Pseudomonas aeruginosa GF31,简称菌株GF31),采用气相色谱/质谱联用(GC/MS)分析技术,开展菌株GF31在实际土壤环境中对氯氰菊酯的降解特性和降解产物研究,并进行了模拟田间实验.结果表明,在土壤中菌株GF31降解氯氰菊酯的主要产物为二氯菊酸和间苯...     

Isolation of mesotrione-degrading bacteria from aquatic environments in Brazil

Mesotrione is a benzoylcyclohexane-1,3-dione herbicide that inhibits 4-hydroxyphenyl pyruvate dioxygenase in target plants. Although it has been used since 2000, only a limited number of degrading microorganisms have been reported. Mesotrione-degrading bacteria were selected among strains isolated from Brazilian aquatic environments, located near corn fields treated with this herbicide. Pantoea ananatis was found to rapidly and completely degrade mesotrione. Mesotrione did not serve as a sole C, N, or S source for growth of P. ananatis, and mesotrione catabolism required glucose supplementation to minimal media. LC-MS/MS analyses indicated that mesotrione degradation produced intermediates other than 2-amino-4-methylsulfonyl benzoic acid or 4-methylsulfonyl-2-nitrobenzoic acid, two metabolites previously identified in a mesotrione-degrading Bacillus strain. Since P. ananatis rapidly degraded mesotrione, this strain might be useful for bioremediation purposes.     

Dimethylphthalate hydrolysis by specific microbial esterase

TWO BACTERIAL STRAINS: Arthrobacter sp. and Sphingomonas paucimobilis were isolated from soil by enrichment cultures using dimethylphthalate (DMP) or monomethylphthalate (MMP) as sole carbon source, respectively. DMP was rapidly transformed by an Arthrobacter sp. culture with formation of MMP and phthalic acid (PA) which is further degraded. This strain was unable to hydrolyse MMP. A mechanism of degradation of DMP was proposed with two ways: DMP-->PA and DMP-->MMP. The S. paucimobilis strain hydrolyses only MMP and a coculture of the two strains allowed a complete degradation of DMP.     

共代谢基质对芽孢杆菌Y-4降解异喹啉的影响

以吡啶，葡萄糖和邻苯二甲酸作为共代谢基质，研究了它们对芽孢杆菌Y_4降解异喹啉的影响。实验结果表明各降解过程均遵循二级反应动力学方程：-dS／dt=K2S2＋K1S＋K0。吡啶的加入会抑制异喹啉的降解，并且吡啶的浓度越高，抑制作用越明显。反应体系中葡萄糖的浓度为100-800mg／L时，葡萄糖的加入会促进异喹啉的降解，且葡萄糖浓度越大，异喹啉降解速率P越大，当葡萄糖的浓度为800mg／L时，其降解率速率P可由未加葡萄糖的0．1924h。上升为0．2255h-1。适宜浓度的邻苯二甲酸会对异喹啉的降解产生促进作用，邻苯二甲酸的浓度为50mg／L时，异喹啉的降解速率可由原来的0．1924h-1增加到0．2145h-1，邻苯二甲酸浓度过高反而会抑制异喹啉的降解。     

Benzoate degradation by Rhodococcus opacus 1CP after dormancy: Characterization of dioxygenases involved in the process

The process of benzoate degradation by strain Rhodococcus opacus 1CP after a five-year dormancy was investigated and its peculiarities were revealed. The strain was shown to be capable of growth on benzoate at a concentration of up to 10 g L^?1. The substrate specificity of benzoate dioxygenase (BDO) during the culture growth on a medium with a low (200–250 mg L^?1) and high (4 g L^?1) concentration of benzoate was assessed. BDO of R. opacus 1CP was shown to be an extremely narrow specificity enzyme. Out of 31 substituted benzoates, only with one, 3-chlorobenzoate, its activity was higher than 9% of that of benzoate. Two dioxygenases, catechol 1,2-dioxygenase (Cat 1,2-DO) and protocatechuate 3,4-dioxygenase (PCA 3,4-DO), were identified in a cell-free extract, purified and characterized. The substrate specificity of Cat 1,2-DO isolated from cells of strain 1CP after the dormancy was found to differ significantly from that of Cat 1,2-DO isolated earlier from cells of this strain grown on benzoate. By its substrate specificity, the described Cat 1,2-DO was close to the Cat 1,2-DO from strain 1CP grown on 4-methylbenzoate. Neither activity nor inhibition by protocatechuate was observed during the reaction of Cat 1,2-DO with catechol, and catechol had no inhibitory effect on the reaction of PCA 3,4-DO with protocatechuate.     

Migration of heavy metals and migration-degradation of phenanthrene in soil using electro kinetic-laccase combined remediation system

Abstract

In order to solve the problem of heavy metal-organic compound soil pollution, in this paper, we developed a highly efficient electro kinetic-laccase combined remediation (EKLCR) system. The results showed that the EKLCR system had an obvious migration effect on heavy metals (copper and cadmium) and good migration-degradation effect on phenanthrene. The migration rates of copper and cadmium were 48.3% and 40.3%, respectively. Especially, with the presence of laccase, the removal rate of phenanthrene on Cu²⁺-contaminated soil was higher than that of Cd²⁺-contaminated soil due to the significant effect of heavy metals on the enzymatic activity of laccase. The average migration-degradation rate of phenanthrene by EKLCR system was 45.4%. Finally, gas chromatography-mass spectrometry (GC/MS) was used to analyze the degradation intermediates of phenanthrene in the soil, which included 9,10-Phenanthrenequinone, phthalic acid, and 2,2-Biphenyldicarboxylic Acid. In addition, we give the possible degradation pathways of phenanthrene, 2,2-Biphenyldicarboxylic Acid is further degraded to produce phthalic acid. The products of the phthalic acid metabolic pathway are protocatechuic acid, pyruvic acid or succinic acid, the final products of these organic acids are carbon dioxide and water.