Cometabolic Transformation Research Articles

Facilitation of Co-Metabolic Transformation and Degradation of Monochlorophenols by Pseudomonas sp. CF600 and Changes in Its Fatty Acid Composition.

In this study, co-metabolic degradation of monochlorophenols (2-CP, 3-CP, and 4-CP) by the Pseudomonas sp. CF600 strain in the presence of phenol, sodium benzoate, and 4-hydroxybenzoic acid as an additional carbon source as well as the survival of bacteria were investigated. Moreover, the changes in cellular fatty acid profiles of bacteria depending on co-metabolic conditions were analyzed. It was found that bacteria were capable of degrading 4-CP completely in the presence of phenol, and in the presence of all substrates, they degraded 2-CP and 3-CP partially. The highest 2-CP and 3-CP removal was observed in the presence of sodium benzoate. Bacteria exhibited three various dioxygenases depending on the type of growth substrate. It was also demonstrated that bacteria exposed to aromatic growth substrates earlier degraded monochlorophenols more effectively than unexposed cells. The analysis of fatty acid profiles of bacteria indicated the essential changes in their composition, involving alterations in fatty acid saturation, hydroxylation, and cyclopropane ring formation. The most significant change in bacteria exposed to sodium benzoate and degrading monochlophenols was the appearance of branched fatty acids. The knowledge from this study indicates that Pseudomonas sp. CF600 could be a suitable candidate for the bioaugmentation of environments contaminated with phenolic compounds.

Co-metabolic transformation of the neonicotinoid insecticide imidacloprid by the new soil isolate Pseudoxanthomonas indica CGMCC 6648

A new imidacloprid (IMI) degrading bacterium Z-9 (deposited number CGMCC 6648) was isolated and identified as Pseudoxanthomonas indica by 16S rRNA gene analysis. Two metabolites were identified as olefin and 5-hydroxy IMI by liquid chromatography-mass spectrometry and nuclear magnetic resonance analysis. P. indica CGMCC 6648 degraded 70.1% of IMI (1.22 mmol L−1) and formed 0.93 mmol L−1 5-hydroxy IMI and 0.05 mmol L−1 olefin IMI in 6 days and in the presence of 100 mmol L−1 glucose. The half-life of IMI degradation was 3.6 days. P. indica CGMCC 6648 transforms IMI via a co-metabolism mechanism and different carbohydrates have significant effects on 5-hydroxy IMI formation, whereas different organic acids have substantial effects on olefin IMI production. Lactose is the best co-substrate for IMI degradation and 5-hydroxy IMI formation with 0.77 mmol L−1 degraded and 0.67 mmol L−1 formed in 48 h, respectively. Pyruvate is the best co-substrate for olefin IMI formation with 0.17 mmol L−1 produced in 96 h for all carbon sources tested. Pyruvate significantly stimulates the conversion of 5-hydroxy IMI to olefin IMI, whereas glucose slightly inhibits this reaction. P. indica CGMCC 6648 rapidly degrades IMI and forms olefin IMI, which may enhance its potential for biodegradation of IMI and increase its insecticidal activity, which can decrease the IMI dosage required.

Potential of Cometabolic Transformation of Polysaccharides and Lignin in Lignocellulose by Soil Actinobacteria

While it is known that several Actinobacteria produce enzymes that decompose polysaccharides or phenolic compounds in dead plant biomass, the occurrence of these traits in the environment remains largely unclear. The aim of this work was to screen isolated actinobacterial strains to explore their ability to produce extracellular enzymes that participate in the degradation of polysaccharides and their ability to cometabolically transform phenolic compounds of various complexities. Actinobacterial strains were isolated from meadow and forest soils and screened for their ability to grow on lignocellulose. The potential to transform 14C-labelled phenolic substrates (dehydrogenation polymer (DHP), lignin and catechol) and to produce a range of extracellular, hydrolytic enzymes was investigated in three strains of Streptomyces spp. that possessed high lignocellulose degrading activity. Isolated strains showed high variation in their ability to produce cellulose- and hemicellulose-degrading enzymes and were able to mineralise up to 1.1% and to solubilise up to 4% of poplar lignin and to mineralise up to 11.4% and to solubilise up to 64% of catechol, while only minimal mineralisation of DHP was observed. The results confirm the potential importance of Actinobacteria in lignocellulose degradation, although it is likely that the decomposition of biopolymers is limited to strains that represent only a minor portion of the entire community, while the range of simple, carbon-containing compounds that serve as sources for actinobacterial growth is relatively wide.

Bioaugmentation with DistinctDehalobacterStrains Achieves Chloroform Detoxification in Microcosms

Chloroform (CF) is a widespread groundwater contaminant not susceptible to aerobic degradation. Under anoxic conditions, CF can undergo abiotic and cometabolic transformation but detoxification is generally not achieved. The recent discovery of distinct Dehalobacter strains that respire CF to dichloromethane (DCM) and ferment DCM to nonchlorinated products promises that bioremediation of CF plumes is feasible. To track both strains, 16S rRNA gene-based qPCR assays specific for either Dehalobacter strain were designed and validated. A laboratory treatability study explored the value of bioaugmentation and biostimulation to achieve CF detoxification using anoxic microcosms established with aquifer material from a CF-contaminated site. Microcosms that received 6% (v/v) of the CF-to-DCM-dechlorinating culture Dhb-CF to achieve an initial Dehalobacter cell titer of 1.6 ± 0.9 × 10(4) mL(-1) dechlorinated CF to stoichiometric amounts of DCM. Subsequent augmentation with 3% (v/v) of the DCM-degrading consortium RM to an initial Dehalobacter cell abundance of 1.2 ± 0.2 × 10(2) mL(-1) achieved complete DCM degradation in microcosms amended with 10 mM bicarbonate. Growth of the CF-respiring and the DCM-degrading Dehalobacter populations and detoxification were also observed in microcosms that received both inocula simultaneously. These findings suggest that anaerobic bioremediation (e.g., bioaugmentation) is a possible remedy at CF- and DCM-contaminated sites without CT, which strongly inhibited CF organohalide respiration and DCM organohalide fermentation.

Using Compound-Specific Isotope Analysis to Assess Biodegradation of Nitroaromatic Explosives in the Subsurface

Assessing the fate of nitroaromatic explosives in the subsurface is challenging because contaminants are present in different phases (e.g., bound to soil or sediment matrix or as solid-phase residues) and transformation takes place via several potentially competing pathways over time-scales of decades. We developed a procedure for compound-specific analysis of stable C, N, and H isotopes in nitroaromatic compounds (NACs) and characterized biodegradation of 2,4,6-trinitrotoluene (TNT) and two dinitrotoluene isomers (2,4-DNT and 2,6-DNT) in subsurface material of a contaminated site. The type and relative contribution of reductive and oxidative pathways to the degradation of the three contaminants was inferred from the combined evaluation of C, N, and H isotope fractionation. Indicative trends of Δδ(15)N vs Δδ(13)C and Δδ(2)H vs Δδ(13)C were obtained from laboratory model systems for biodegradation pathways initiated via (i) dioxygenation, (ii) reduction, and (iii) CH3-group oxidation. The combined evaluation of NAC isotope fractionation in subsurface materials and in laboratory experiments suggests that in the field, 86-89% of 2,4-DNT transformation was due to dioxygenation while TNT was mostly reduced and 2,6-DNT reacted via a combination of reduction and CH3-group oxidation. Based on historic information on site operation, our data imply biodegradation of 2,4-DNT with half-lives of up to 9-17 years compared to 18-34 years for cometabolic transformation of TNT and 2,6-DNT.

Co-metabolic degradation of chlorobenzene by the fluorobenzene degrading wild strain Labrys portucalensis

Due to the widespread use of chlorobenzene (CB), environmental contamination with this compound is a major concern. A previously isolated bacterium named Labrys portucalensis (strain F11), that is able to use fluorobenzene (FB) as sole carbon and energy source, was tested for its capability to degrade CB. Strain F11 was able to partially degrade CB only when F11 cells were previously grown in FB. Biodegradation of 0.5 mM of CB was achieved at a rate of 7.95 ± 0.39 μmol l−1 day−1 with concomitant stoichiometric release of 50% of the chloride, while degradation of 1 mM of this compound resulted in 85% degradation at a rate of 16.9 ± 0.81 μmol l−1 day−1 and with a 15% chloride release on the basis of the amount of compound biodegraded. Total CB biodegradation and dechlorination was only achieved when FB was also supplied to F11 cultures, suggesting cometabolic transformation. Total degradation of 0.5 mM of CB and 0.5 mM of FB occurred simultaneously at degradation rates of 105 ± 6.07 μmol l−1 day−1 and 126 ± 16.2 μmol l−1 day−1 respectively, with stoichiometric halogen release. Growth yield was lower when both substrates were present, suggesting synergistic inhibition. To our knowledge, this is the first time that co-metabolic biodegradation of CB in the presence of the fluorinated analogue is reported.

Proteogenomic evidence for β-oxidation of plant-derived 3-phenylpropanoids in “Aromatoleum aromaticum ” EbN1

The betaproteobacterium "Aromatoleum aromaticum" EbN1 utilizes eight different plant-derived nonhydroxylated (e.g. cinnamate) and hydroxylated (e.g. p-coumarate) 3-phenylpropanoids with nitrate as electron acceptor. Differential protein profiling (2D-DIGE) revealed abundance increases of five proteins (EbA5316 to EbA5320) during anaerobic growth with cinnamate, hydrocinnamate, p-coumarate, and 3-(4-hydroxyphenyl)propanoate, compared to anaerobic benzoate-adapted cells serving as reference state. The predicted functions of four of these proteins (EbA5317, fatty acid-coenzyme A (CoA) ligase; EbA5318, enoyl-CoA hydratase/isomerase; EbA5319, β-ketothiolase; and EbA5320, 3-hydroxyacyl-CoA dehydrogenase) suggest β-oxidation of the above 3-phenylpropanoids to benzoyl-CoA and p-hydroxybenzoyl-CoA, respectively. The fifth protein (EbA5316, ABC-type periplasmic solute-binding protein) could be involved in 3-phenylpropanoid uptake. The detection of 3-hydroxy-3-phenylpropanoate during anaerobic growth with cinnamate and hydrocinnamate or 3-hydroxy-3-(4-hydroxyphenyl)propanoate during anaerobic growth with p-coumarate and 3-(4-hydroxyphenyl)propanoate supports the proteome-predicted β-oxidation pathway. Based on the specific formation of EbA5316-20 also during anaerobic growth with further 3-phenylpropanoid growth substrates including cinnamyl alcohol, m-coumarate, 3-(3,4-dihydroxyphenyl)propanoate and 3,4-dihydroxycinnamate (caffeate), a common β-oxidation route is proposed for 3-phenylpropanoid degradation in strain EbN1. The low amount of metabolites attributable to cometabolic transformation of nongrowth supporting 3-phenylpropanoids (e.g. o-coumarate, ferulate) may be indicative for a high substrate specificity of the involved enzymes.

Effect of S0/X0 ratio and acclimation on respirometry of activated sludge in the cometabolic biodegradation of phenolic compounds

Aerobic batch biodegradation experiments and respirometric analysis were performed in order to investigate the effect of S0/X0 (substrate/biomass) ratio and preliminary acclimation on bi-solute biodegradation of phenolic compounds. It was shown that 2-chlorophenol (2-CP) and 2-nitrophenol (2-NP) could be cometabolically biodegraded only with acclimated biomass in the presence of phenol as growth substrate. Acclimation resulted in domination of phenol oxidizing bacteria which could induce the necessary enzymes for cometabolic transformation of 2-CP and 2-NP. Biodegradation of the cometabolic compounds occurred even after depletion of phenol at resting cell conditions. Both compounds could be successfully biodegraded by the acclimated biomass at initial substrate concentrations as high as 300mg/L. Respirometric analysis showed that the optimum S0/X0 ratio ranged between 1.5 and 5.5mg CODeq/mg MLSS for cometabolic substrates 2-CP and 2-NP, whereas it was as high as 8.5mg CODeq/mg MLSS for phenol which corresponds to a phenol concentration of about 1500mg/L.

Reductive dehalogenation mediated initiation of aerobic degradation of 2-chloro-4-nitrophenol (2C4NP) by Burkholderia sp. strain SJ98

Burkholderia sp. strain SJ98 (DSM 23195) was previously isolated and characterized for degradation and co-metabolic transformation of a number nitroaromatic compounds. In the present study, we evaluated its metabolic activity on chlorinated nitroaromatic compounds (CNACs). Results obtained during this study revealed that strain SJ98 can degrade 2-chloro-4-nitrophenol (2C4NP) and utilize it as sole source of carbon, nitrogen, and energy under aerobic conditions. The cells of strain SJ98 removed 2C4NP from the growth medium with sequential release of nearly stoichiometric amounts of chloride and nitrite in culture supernatant. Under aerobic degradation conditions, 2C4NP was transformed into the first intermediate that was identified as p-nitrophenol by high-performance liquid chromatography, LCMS-TOF, and GC-MS analyses. This transformation clearly establishes that the degradation of 2C4NP by strain SJ98 is initiated by "reductive dehalogenation"; an initiation mechanism that has not been previously reported for microbial degradation of CNAC under aerobic conditions.

Aerobic cometabolism of trichloroethene and cis-dichloroethene with benzene and chlorinated benzenes as growth substrates

Using inoculum from a microcosm study that exhibited aerobic transformation of cis-1,2-dichloroethene (cDCE) and trichloroethene (TCE) commensurate with biodegradation of monoaromatic compounds, enrichment cultures were developed by providing benzene, chlorobenzene (CB), dichlorobenzene (DCB) isomers and 1,2,4-trichlorobenzene as carbon and energy sources. Isolates that grow on benzene, CB, 1,2-DCB and 1,3-DCB were identified as Rhodococcus, Ralstonia, Variovorax and Ralstonia spp., respectively. Cometabolic transformation of cDCE and TCE by resting cells was demonstrated. Transformation capacities ( T c = 0.47–1.0 μg TCE mg −1 biomass; 1.3–5.3 μg cDCE mg −1 biomass), transformation yields ( T y = 0.18-0.27 μg TCE mg −1 substrate; 0.46–2.1 μg cDCE mg −1 substrate), and pseudo-first-order cometabolic degradation rate constants (0.00081–0.0031 L mg TCE −1 d −1; 0.0012–0.030 L mg cDCE −1 d −1) for resting cells grown on benzene, CB, 1,2-DCB and 1,3-DCB were generally lower in comparison to phenol and toluene-grown isolates. Cometabolic transformation of cDCE and TCE also occurred while the cultures concurrently consumed their growth substrate ( T c ′ = 0.15–0.33 μg TCE mg −1 biomass; 4.9–11 μg cDCE mg −1 biomass; T y ′ = 0.06–0.11 μg TCE mg −1 substrate; 1.7–4.6 μg cDCE mg −1 substrate), a condition that is more likely to be encountered in situ compared to cometabolic transformations by resting cells. This study is the first to report transformation rates, capacities, and yields for cometabolism of cDCE and TCE during aerobic growth on benzene, CB, 1,2-DCB and 1,3-DCB. This type of information is needed to predict the potential for natural attenuation when these compounds occur as co-contaminants.

Enhancement of cometabolic biodegradation of 4-chlorophenol induced with phenol and glucose as carbon sources by Comamonas testosteroni

The biological degradation of phenol and 4-chlorophenol (4-CP) by Comamonas testosteroni CECT 326T has been studied. Phenol and 4-CP were treated alone as a sole carbon and energy source, but only phenol was completely degraded by C. testosteroni. Since the presence of cosubstrates can enhance the toxic compounds removal by pure cultures, phenol and glucose were added as growth substrates for cometabolic transformation of 4-CP. High efficiencies were obtained in all the experiments carried out in presence of both cosubstrates. In spite of the fact that the addition of glucose reduced the lag phase of 4-CP removal, lower phenol concentrations were required to obtain the same degradation efficiencies. The cometabolic transformation of 4-CP was closely related with the extent of phenol removal. The values of the 4-CP/biomass concentration ratio (S/X) obtained for discriminating between complete (S/X ≤ 0.11) and partial 4-CP (S/X ≥ 0.31) transformation showed a narrower range than that reported in the literature. The extent of the cometabolic 4-CP transformation in the presence of phenol could be further enhanced by using glucose as an additional carbon and energy source. However, no significant influence of glucose concentration on 4-CP removal was observed over the concentration range studied.

The role of organic matter in the removal of emerging trace organic chemicals during managed aquifer recharge

This study explored the effect of different bulk organic carbon matrices on the fate of trace organic chemicals (TOrC) during managed aquifer recharge (MAR). Infiltration through porous media was simulated in biologically active column experiments under aerobic and anoxic recharge conditions. Wastewater effluent derived organic carbon types, differing in hydrophobicity and biodegradability (i. e., hydrophobic acids, hydrophilic carbon, organic colloids), were used as feed substrates in the column experiments. These carbon substrates while fed at the same concentration differed in their ability to support soil biomass growth during porous media infiltration. Removal of degradable TOrC (with the exception of diclofenac and propyphenazone) was equal or better under aerobic versus anoxic porous media infiltration conditions. During the initial phase of infiltration, the presence of biodegradable organic carbon (BDOC) enhanced the decay of degradable TOrC by promoting soil biomass growth, suggesting that BDOC served as a co-substrate in a co-metabolic transformation of these contaminants. However, unexpected high removal efficiencies were observed for all degradable TOrC in the presence of low BDOC concentrations under well adopted oligotrophic conditions. It is hypothesized that removal under these conditions is caused by a specialized microbial community growing on refractory carbon substrates such as hydrophobic acids. Findings of this study reveal that the concentration and character of bulk organic carbon present in effluents affect the degradation efficiency for TOrC during recharge operation. Specifically aerobic, oligotrophic microbiological soil environments present favorable conditions for the transformation of TOrC, including rather recalcitrant compounds such as chlorinated flame retardants.

Physiological comparison of Pseudomonas putida between two growth phases during cometabolism of 4‐chlorophenol in presence of phenol and glutamate: a proteomics approach

AbstractBACKGROUND: In cometabolic transformation of 4‐chlorophenol (4‐cp) in the presence of phenol and sodium glutamate (SG), a new biphasic growth pattern has been reported. This study investigates the physiological changes of Pseudomonas putida P8 during the biphasic growth by means of 2‐dimensional gel electrophoresis (2‐DE) and MALDI‐TOF analyses.RESULTS: A total of 49 protein spots were selected and identified in the 2‐DE gels from P. putida P8 grown on a mixed substrate containing 200 mg L−1 of phenol, 200 mg L−1 of 4‐cp and 1000 mg L−1 SG. Among them, 16 protein spots were found differentially expressed in the two exponential growth phases during the biphasic growth, including six catabolic enzymes (DmpC, DmpD, DmpE, DmpF, DmpG and AspA) for substrate utilization. The expression of other proteins involved in detoxification and stress responses, carbohydrate and energy metabolism, and environmental information processing as well as a multifunctional xenobiotic reductase (XenA) was quantitatively analyzed and discussed.CONCLUSION: The expression levels of the identified catabolic enzymes during growth in the two growth phases correlated well with the substrate utilization patterns observed in previous kinetics studies. Furthermore, the results show that cells growing on a mixture of aromatic substrates undergo significant physiological changes. Copyright © 2009 Society of Chemical Industry

Microbial transformation of benzothiophenes, with carbazole as the auxiliary substrate, by Sphingomonas sp. strain XLDN2-5

Benzothiophenes are a toxic and relatively recalcitrant fraction of coal-tar creosote. We investigated the co-metabolic transformation of benzothiophene (BT) and its derivatives by the carbazole (CA) degrader Sphingomonas sp. XLDN2-5, which is not able to grow on benzothiophenes as the sole carbon source. Among the benzothiophenes tested, BT, 2-methylbenzothiophene (2-MBT) and 5-methylbenzothiophene (5-MBT) were co-metabolically converted. For 3-methylbenzothiophene, there was complete inhibition of growth on CA. The common transformation products for BT, 2-MBT and 5-MBT are the corresponding sulfoxides and sulfones. For BT, several high-molecular-mass sulfur-containing aromatic compounds, including benzo[b]naphtho[1,2-d]thiophene, benzo[b]naphtho[1,2-d]thiophene-7-oxide, 6a,11b-dihydrobenzo[b]naphtho[1,2-d]thiophene, 6a,11b-dihydrobenzo[b]naphtho[1,2-d]thiophene-7-oxide, and a new product, 6,12-epithiobenzo[b]naphtho[1,2-d]thiophene, were detected by GC-MS. These high-molecular-mass products are thought to be generated from a Diels-Alder-type reaction. Investigations with a combination of GC and flame ionization detection showed that about 17 % of BT was transformed to benzo[b]naphtho[1,2-d]thiophene. Aerobic transformation of benzothiophenes to sulfoxides and sulfones can reduce their toxicity, and facilitate their biodegradation. However, the formation of the high-molecular-mass products, such as benzo[b]naphtho[1,2-d]thiophene, should be considered in the biodegradation of benzothiophenes.

Co-metabolism of di- and trichlorobenzoates in a 2-chlorobenzoate-degrading bacterial culture: Effect of the position and number of halo-substituents

The degradative properties towards chlorobenzoates by a 2-chlorobenzoate-degrading mixed culture (2MC) were investigated. Although 2MC did not grow on 2,3,5-; 2,3,6- and 2,4,6-trichlorobenzoates, it was able to completely oxidise 2,3,5-trichlorobenzoate and 2,5-dichlorobenzoate. This was degraded through the intermediate formation of 4-chlorocatechol and 3- cis, cis-chloromuconate, suggesting a dioxygenation in 1,2 position. 4-Chlorobenzoate; 3,4- and 2,6-dichlorobenzoate; 2,4,6- and 2,3,6-trichlorobenzoate did not undergo any co-metabolic transformation, the only 2,4-dichlorobenzoate was scarcely transformed into a dichlorinated phenol as end-product. 2MC was constituted of three prominent species: Stenotrophomonas maltophilia, Cupriavidus necator and Flavobacterium sp. Ring-hydroxylating dioxygenase genes ( BenA) were retrieved in S. maltophilia and C. necator, indicating the presence of a “modified ortho pathway” in the culture. On the basis of its degradative properties, 2MC could be a good candidate for use in the removal of ortho- and/or meta-chlorobenzoates that are the major end-products of PCB co-metabolism.

Degradation and mineralization of 2-chloro-, 3-chloro-and 4-chlorobiphenylby a newly characterized natural bacterial strain isolated froman electrical transformer fluid-contaminated soil

A bacterium classified as Achromobacter xylosoxidans strain IR08 by phenotypic typing coupled with 16S rRNA gene analysis was isolated from a soil contaminated with electrical transformer fluid for over sixty years using Aroclor 1221 as an enrichment substrate. The substrate utilization profiles revealed that IR08 could grow on all three monochlorobiphenyls (CBs), 2,4'- and 4,4'-dichlorobiphenyl as well as 2-chlorobenzoate (2-CBA), 3-CBA, 4-CBA, and 2,3-dichlorobenzoate. Unusually, growth was poorly sustained on biphenyl and benzoate. In growth experiments, IR08 degraded all CBs (0.27 mmol/L) in less than 96 h with concomitant stoichiometric release of inorganic chloride and growth yields were 2-3 times higher than those observed on biphenyl. In contrast to most of the chlorobiphenyl-degrading strains described in the literature, which are reported to form CBA, no metabolite was identified in the culture broth by HPLC analysis. When co-incubated with respective CBs and biphenyl, strain IR08 preferentially utilized the chlorinated analogues in less than 96 h while it took another 264 h before 90% of the initially supplied biphenyl could be degraded. The promotion of co-metabolic transformation of halogenated substrates by the inclusion of their non-halogenated derivatives may not therefore, result in universal benefits.

Utilization of Fluoroethene as a Surrogate for Aerobic Vinyl Chloride Transformation

Fluoroethene (FE) is a stable molecule in aqueous solution and its aerobic transformation potentially yields F-. This work evaluated if FE is a suitable surrogate for monitoring aerobic vinyl chloride (VC) utilization or cometabolic transformation. Experiments were carried out with three isolates, Mycobacterium strain EE13a, Mycobacterium strain JS60, and Nocardioides strain JS614 to evaluate if their affinities for FE and VC and their rates of transformation were comparable and whether the transformation of FE and F- accumulation could be correlated with VC utilization. JS614 grew on FE in addition to VC, making it the first organism reported to use FE as a sole carbon and energy source. EE13a cometabolized VC and FE, and JS60 catabolized VC and cometabolized FE. There was little difference among the three strains in the Ks or kmax values for VC or FE. Competitive inhibition modeled the temporal responses of FE and VC transformations and Cl- and F- release when both substrates were present. Both the rate of FE transformation and rate of F-accumulation could be correlated with the rate of aerobic transformation of VC and showed promise for estimating VC rates in situ using FE as a reactive surrogate.

Hybrid-Hollow-Fiber Membrane Bioreactor for Cometabolic Transformation of 4-Chlorophenol in the Presence of Phenol

Hybrid-hollow-fiber membrane bioreactors were developed for the enhanced cometabolic biotransformation of phenol and 4-chlorophenol (4-cp) by Pseudomonas putida ATCC49451. Bioreactor performance was investigated, compared, and analyzed under batch and continuous operating modes. The spinning solutions contained polysulfone (PS), N -methyl-2-pyrrolidone, and various weight ratios of granular activated carbon (GAC) (GAC: PS of 0, 1:4, and 1:2). The bioreactor fabricated with 1:2 GAC hybrid-hollow-fiber membranes demonstrated the best performance for the removal of phenol and 4-cp, both under batch and continuous operations. Under batch operation, 500 mg L−1 phenol and 4-cp were completely removed within 23 h in the bioreactor, compared with 26 and 30 h for the 1:4 GAC and GAC free bioreactors. Sorption, biotransformation, desorption, and bioregeneration were identified as the four steps for substrate removal during batch operation. The 1:2 GAC hollow-fiber membrane bioreactor also manifested superiority ove...

Biodegradation kinetics of the nitramine explosive CL-20 in soil and microbial cultures

The cyclic nitramine explosive CL-20 (C(6)H(6)N(12)O(12), 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12 -hexaazaisowurtzitane) is a relatively new energetic compound which could be a persistent organic pollutant. To follow its biodegradation dynamics, CL-20 was added to soil alone or together with organic co-substrates and N-source and incubated under oxic and anoxic conditions. Without co-substrates, the CL-20 degradation was detectable only under anoxic conditions. The highest degradation rate was found under aerobic conditions and with the addition of co-substrates, succinate and pyruvate being more efficient than acetate, glucose, starch or yeast extract. When added to intact soil, CL-20 degradation was not affected by the N content, but in soil serially diluted with N-free succinate-mineral medium, the process became N-limited. About 40% of randomly selected bacterial colonies grown on succinate agar medium were able to decompose CL-20. Based on 16S rDNA gene sequence and cell morphology, they were affiliated to Pseudomonas, Rhodococcus, Ochrobactrum, Mycobacterium and Ralstonia. In the pure culture of Pseudomonas sp. MS-P grown on the succinate-mineral N(+) medium, the degradation kinetics were first order with the same apparent kinetic constant throughout growth and decline phases of the batch culture. The observed kinetics agreed with the model that supposes co-metabolic transformation of CL-20 uncoupled from cell growth, which can be carried out by several constitutive cellular enzymes with wide substrate specificity.

Kinetics of 1,4-Dioxane Biodegradation by Monooxygenase-Expressing Bacteria

1,4-Dioxane is a probable human carcinogen, and an important emerging water contaminant. In this study, the biodegradation of dioxane by 20 bacterial isolates was evaluated, and 13 were found to be capable of transforming dioxane. Dioxane served as a growth substrate for Pseudonocardia dioxanivorans CB1190 and Pseudonocardia benzenivorans B5, with yields of 0.09 g protein g dioxane(-1) and 0.03 g protein g dioxane(-1), respectively. Cometabolic transformation of dioxane was observed for monooxygenase-expressing strains that were induced with methane, propane, tetrahydrofuran, or toluene including Methylosinus trichosporium OB3b, Mycobacterium vaccae JOB5, Pseudonocardia K1, Pseudomonas mendocina KR1, Ralstonia pickettii PKO1, Burkholderia cepacia G4, and Rhodococcus RR1. Product toxicity resulted in incomplete dioxane degradation for many of the cometabolic reactions. Brief exposure to acetylene, a known monooxygenase inhibitor, prevented oxidation of dioxane in all cases, supporting the hypothesis that monooxygenase enzymes participated in the transformation of dioxane by these strains. Further, Escherichia coli TG1/pBS(Kan) containing recombinant plasmids derived from the toluene-2- and toluene-4-monooxygenases of G4, KR1 and PKO1 were also capable of cometabolic dioxane transformation. Dioxane oxidation rates measured at 50 mg/L ranged from 0.01 to 0.19 mg hr(-1) mg protein(-1) for the metabolic processes, 0.1-0.38 mg hr(-1) mg protein(-1) for cometabolism by the monooxygenase-induced strains, and 0.17-0.60 mg hr(-1) mg protein(-1) for the recombinant strains. Dioxane was not degraded by M. trichosporium OB3b expressing particulate methane monooxygenase, Pseudomonas putida mt-2 expressing a toluene side-chain monooxygenase, and PseudomonasJS150 and Pseudomonas putida F1 expressing toluene-2,3-dioxygenases. This is the first study to definitively show the role of monooxygenases in dioxane degradation using several independent lines of evidence and to describe the kinetics of metabolic and cometabolic dioxane degradation.