31 results on '"Koutinas M"'
Search Results
2. Improvement of anaerobic digestion process to reduce ammonia production
- Author
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Montoneri, E., Padoan, E., Negre, M., Boero, V., Solaro, S., Antonini, M., Kallis, M. Andreou K., Vyrides, I., Koutinas, M., Photiou, P., Konstantinidis, V., Riggio, V., Negro, V., and Mainero, D.
- Published
- 2019
3. A NEW ANAEROBIC FERMENTATION PROCESS TO PRODUCE DIGESTATE WITH LOW AMMONIUM CONTENT
- Author
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Montoneri, E., Fabbri, Gloria, Negre, M., Boero, V., Solaro, S., Antonini, M., Kallis, M., Andreou, K., Vyrides, I., Koutinas, M., Photiou, P., Konstantinidis, V., Riggio, V., Negro, V., and Mainero, D.
- Published
- 2018
4. High temperature alcoholic fermentation of orange peel by the newly isolated thermotolerant Pichia kudriavzeviiKVMP10
- Author
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Koutinas, M., primary, Patsalou, M., additional, Stavrinou, S., additional, and Vyrides, I., additional
- Published
- 2015
- Full Text
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5. The regulatory logic of m-xylene biodegradation by Pseudomonas putida mt-2 exposed by dynamic modelling of the principal node Ps/Pr of the TOL plasmid
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Koutinas, M., Lam, M.C., Kiparissides, A., Silva-Rocha, R., Godinho, M., Livingston, A.G., Pistikopoulos, E.N., de Lorenzo, V., Martins Dos Santos, V.A.P., and Mantalaris, A.
- Subjects
global sensitivity-analysis ,Systeem en Synthetische Biologie ,Bacteria ,Pseudomonas putida ,pathway ,translation ,Biological Sciences ,pu promoter ,escherichia-coli ,Systems and Synthetic Biology ,activation ,xylr ,Natural Sciences ,transcription ,Catabolite repression ,catabolite repression ,rna-polymerase - Abstract
P>The structure of the extant transcriptional control network of the TOL plasmid pWW0 born by Pseudomonas putida mt-2 for biodegradation of m-xylene is far more complex than one would consider necessary from a mere engineering point of view. In order to penetrate the underlying logic of such a network, which controls a major environmental cleanup bioprocess, we have developed a dynamic model of the key regulatory node formed by the Ps/Pr promoters of pWW0, where the clustering of control elements is maximal. The model layout was validated with batch cultures estimating parameter values and its predictive capability was confirmed with independent sets of experimental data. The model revealed how regulatory outputs originated in the divergent and overlapping Ps/Pr segment, which expresses the transcription factors XylS and XylR respectively, are computed into distinct instructions to the upper and lower catabolic xyl operons for either simultaneous or stepwise consumption of m-xylene and/or succinate. In this respect, the model reveals that the architecture of the Ps/Pr is poised to discriminate the abundance of alternative and competing C sources, in particular m-xylene versus succinate. The proposed framework provides a first systemic understanding of the causality and connectivity of the regulatory elements that shape this exemplary regulatory network, facilitating the use of model analysis towards genetic circuit optimization.
- Published
- 2010
6. Revealing microbial strain dynamics case study in an oil-absorber bioscrubber system
- Author
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Baptista, I. R., Koutinas, M., Mantalaris, A., and Andrew Livingston
7. Prediction of double substrate microbial growth kinetics through transcriptional regulation: An integrated experimental/modelling approach on Pseudomonas putida mt-2
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Tsipa, A., Koutinas, M., Stratos Pistikopoulos, Mantalaris, A., and Κουτίνας, Μιχάλης
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Mt-2 ,Pseudomonas putida ,Earth and Related Environmental Sciences ,Natural Sciences
8. Compost stream as a potential biomass for humic acid production: Focus on compost seasonal and geographical variability
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Negro, V., Montoneri, E., Negre, M., Fabbri, G., Boero, V., Simone Solaro, Antonini, M., Koutinas, A., Koutinas, M., Vlysidis, A., Konstantinidis, V., and Mainero, D.
- Subjects
Agricultural Sciences ,Hydrolysis ,Humic acids ,Bio-stimulants ,Compost ,Agriculture Forestry and Fisheries - Abstract
Compost is a voluminous stream rich in humic and fulvic acids, which may be recovered as high-added value compounds. These soluble bio-based lignin-like polymeric substances (SBO) can be extracted through a completely green process developed at pilot scale, whose main core is the hydrolytic route in aqueous solutions at relatively mild temperature (< 140 °C) at ACEA Pinerolese Industriale premises. Due to their chemical-physical properties, the SBO compounds can be used with advantage for myriads of industrial applications, from the formulation of detergents to the production of agriculture biostimulants, answering the increasing demand for bio-compound utilization. In view of LIFECAB project (LIFE16 ENV/IT/000179), the characterization of starting materials and the derived compost has been performed over four seasons and over three European countries (Italy, Greece ad Cyprus). In view of establishing a relationship between SBO molecules and compost properties, this work is a challenging opportunity for assessing the compost variability and its temporal evolution during the composting process. Analyses of pH, salinity, total carbon, total nitrogen and C/N ratio, critically assessed by means of a statistical approach, provide important information about compost composition according to the season and to the local environmental conditions.
9. Microbial dynamics and bioreactor stability in an oil-absorber-bioscrubber system exposed to an alternating sequence of 1,2-dichloroethane and fluorobenzene
- Author
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Koutinas, M., Baptista, I., Meniconi, A., Peeva, L. G., Castro, P. M. L., and Andrew Livingston
10. Ecotoxicological assessment of biomass-derived furan platform chemicals using aquatic and terrestrial bioassays.
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Parmaki S, Vasquez MI, Patsalou M, Gomes RFA, Simeonov SP, Afonso CAM, and Koutinas M
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- Lepidium sativum drug effects, Lepidium sativum growth & development, Ecotoxicology methods, Biological Assay methods, Water Pollutants, Chemical toxicity, Water Pollutants, Chemical analysis, Toxicity Tests methods, Sinapis drug effects, Microalgae drug effects, Furans toxicity, Aliivibrio fischeri drug effects, Biomass
- Abstract
An environmental toxicological assessment of fourteen furanic compounds serving as valuable building blocks produced from biomass was performed. The molecules selected included well studied compounds serving as control examples to compare the toxicity exerted against a variety of highly novel furans which have been additionally targeted as potential or current alternatives to biofuels, building blocks and polymer monomers. The impact of the furan platform chemicals targeted on widely applied ecotoxicity model organisms was determined employing the marine bioluminescent bacterium Aliivibrio fischeri and the freshwater green microalgae Raphidocelis subcapitata , while their ecotoxicity effects on plants were assessed using dicotyledonous plants Sinapis alba and Lepidium sativum . Regarding the specific endpoints evaluated, the furans tested were slightly toxic or practically nontoxic for A. fischeri following 5 and 15 min of exposure. Moreover, most of the building blocks did not affect the growth of L. sativum and S. alba at 150 mg L
-1 for 72 h of exposure. Specifically, 9 and 11 out of the 14 furan platform chemicals tested were non-effective or stimulant for L. sativum and S. alba respectively. Given that furans comprise common inhibitors in biorefinery fermentations, the growth inhibition of the specific building blocks was studied using the industrial workhorse yeast Saccharomyces cerevisiae , demonstrating insignificant inhibition on eukaryotic cell growth following 6, 12 and 16 h of exposure at a concentration of 500 mg L-1 . The study provides baseline information to unravel the ecotoxic effects and to confirm the green aspects of a range of versatile biobased platform molecules.- Published
- 2024
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11. Evolution of prokaryotic colonisation of greenhouse plastics discarded into the environment.
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Martínez-Campos S, Pissaridou P, Drakou K, Shammas C, Andreou K, González-Pleiter M, Fernández-Piñas F, Leganes F, Rosal R, Koutinas M, Kapnisis K, and Vasquez MI
- Subjects
- Bacteria genetics, RNA, Ribosomal, 16S genetics, Rivers, Plastics, Seawater microbiology
- Abstract
Current knowledge on the capacity of plastics as vectors of microorganisms and their ability to transfer microorganisms between different habitats (i.e. air, soil and river) is limited. The objective of this study was to characterise the evolution of the bacterial community adhered to environmental plastics [low-density polyethylene (LDPE)] across different environments from their point of use to their receiving environment destination in the sea. The study took place in a typical Mediterranean intermittent river basin in Larnaka, Cyprus, characterised by a large greenhouse area whose plastic debris may end up in the sea due to mismanagement. Five locations were selected to represent the environmental fate of greenhouse plastics from their use, through their abandonment in soil and subsequent transport to the river and the sea, taking samples of plastics and the surrounding environments (soil and water). The bacterial community associated with each sample was studied by 16S rRNA metabarcoding; also, the main physicochemical parameters in each environmental compartment were analysed to understand these changes. The identification and chemical changes in greenhouse plastics were tracked using Attenuated Total Reflection Fourier Transform Infra-red spectroscopy (ATR-FTIR). Scanning Electron Microscope (SEM) analysis demonstrated an evolution of the biofilm at each sampling location. β-diversity studies showed that the bacterial community adhered to plastics was significantly different from that of the surrounding environment only in samples taken from aqueous environments (freshwater and sea) (p-value p-value > 0.05). The environmental parameters (pH, salinity, total nitrogen and total phosphorus) explained the differences observed at each location to a limited extent. Furthermore, bacterial community differences among samples were lower in plastics collected from the soil than in plastics taken from rivers and seawater. Six genera (Flavobacterium, Altererythrobacter, Acinetobacter, Pleurocapsa, Georgfuchsia and Rhodococcus) were detected in the plastic, irrespective of the sampling location, confirming that greenhouse plastics can act as possible vectors of microorganisms between different environments: from their point of use, through a river system to the final coastal receiving environment. In conclusion, this study confirms the ability of greenhouse plastics to transport bacteria, including pathogens, between different environments. Future studies should evaluate these risks by performing complete sequencing metagenomics to decipher the functions of the plastisphere., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2022
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12. Ultrasound-assisted dilute acid hydrolysis for production of essential oils, pectin and bacterial cellulose via a citrus processing waste biorefinery.
- Author
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Karanicola P, Patsalou M, Stergiou PY, Kavallieratou A, Evripidou N, Christou P, Panagiotou G, Damianou C, Papamichael EM, and Koutinas M
- Subjects
- Cellulose, Hydrolysis, Pectins, Citrus, Oils, Volatile
- Abstract
An orange peel waste biorefinery was developed employing a design of experiments approach to optimize the ultrasound-assisted dilute acid hydrolysis process applied for production of useful commodities. Central composite design-based response surface methodology was used to approximate the combined effects of process parameters in simultaneous production of essential oils, pectin and a sugar-rich hydrolyzate. Application of a desirability function determined the optimal conditions required for maximal production efficiency of essential oils, pectin and sugars as 5.75% solid loading, 1.21% acid concentration and 34.2 min duration. Maximum production yields of 0.12% w/w essential oils, 45% w/w pectin and 40% w/w sugars were achieved under optimized conditions in lab- and pilot-scale facilities. The hydrolyzate formed was applied in bacterial cellulose fermentations producing 5.82 g biopolymer per 100 g waste. Design of experiments was efficient for process analysis and optimization providing a systems platform for the study of biomass-based biorefineries., (Copyright © 2021 Elsevier Ltd. All rights reserved.)
- Published
- 2021
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13. Enhanced biodegradation and valorization of drilling wastewater via simultaneous production of biosurfactants and polyhydroxyalkanoates by Pseudomonas citronellolis SJTE-3.
- Author
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Koutinas M, Kyriakou M, Andreou K, Hadjicharalambous M, Kaliviotis E, Pasias D, Kazamias G, Varavvas C, and Vyrides I
- Subjects
- Biodegradation, Environmental, Pseudomonas, Surface-Active Agents, Wastewater, Polyhydroxyalkanoates
- Abstract
Pseudomonas citronellolis SJTE-3 was isolated as a highly efficient microorganism for biodegradation and valorization of drilling fluids (DF) wastewater. The strain metabolised DF and oily mud exhibiting up to 93%, 86%, 85% and 88% of chemical oxygen demand (COD), n-dodecane, n-tetradecane and naphthalene removal efficiency respectively. Enhanced bioconversion was enabled through production of biosurfactants that reduced the surface tension of water by 53% and resulted in 43.3% emulsification index (E24), while synthesizing 24% of dry cell weight (DCW) as medium-chain-length polyhydroxyalkanoates (PHA). Expression from the main pathways for alkanes and naphthalene biodegradation as well as biosurfactants and PHA biosynthesis revealed that although the alkanes and naphthalene biodegradation routes were actively expressed even at stationary phase, PHA production was stimulated at late stationary phase and putisolvin could comprise the biosurfactant synthesized. The bioconversion of toxic petrochemical residues to added-value thermoelastomers and biosurfactants indicate the high industrial significance of P. citronellolis SJTE-3., (Copyright © 2021 Elsevier Ltd. All rights reserved.)
- Published
- 2021
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14. Immune reconstitution and clinical recovery following anti-CD28 antibody (TGN1412)-induced cytokine storm.
- Author
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Panoskaltsis N, McCarthy NE, Stagg AJ, Mummery CJ, Husni M, Arebi N, Greenstein D, Price CL, Al-Hassi HO, Koutinas M, Mantalaris A, and Knight SC
- Subjects
- Adult, Antibodies, Monoclonal, Humanized pharmacology, Cognitive Dysfunction etiology, Cohort Studies, Cytokine Release Syndrome etiology, Follow-Up Studies, Humans, Male, Young Adult, Antibodies, Monoclonal, Humanized adverse effects, CD28 Antigens agonists, COVID-19 immunology, Cognitive Dysfunction immunology, Cytokine Release Syndrome immunology, Drug-Related Side Effects and Adverse Reactions immunology, Immunotherapy adverse effects, SARS-CoV-2 physiology, T-Lymphocytes immunology
- Abstract
Cytokine storm can result from cancer immunotherapy or certain infections, including COVID-19. Though short-term immune-related adverse events are routinely described, longer-term immune consequences and sequential immune monitoring are not as well defined. In 2006, six healthy volunteers received TGN1412, a CD28 superagonist antibody, in a first-in-man clinical trial and suffered from cytokine storm. After the initial cytokine release, antibody effect-specific immune monitoring started on Day + 10 and consisted mainly of evaluation of dendritic cell and T-cell subsets and 15 serum cytokines at 21 time-points over 2 years. All patients developed problems with concentration and memory; three patients were diagnosed with mild-to-moderate depression. Mild neutropenia and autoantibody production was observed intermittently. One patient suffered from peripheral dry gangrene, required amputations, and had persistent Raynaud's phenomenon. Gastrointestinal irritability was noted in three patients and coincided with elevated γδT-cells. One had pruritus associated with elevated IgE levels, also found in three other asymptomatic patients. Dendritic cells, initially undetectable, rose to normal within a month. Naïve CD8
+ T-cells were maintained at high levels, whereas naïve CD4+ and memory CD4+ and CD8+ T-cells started high but declined over 2 years. T-regulatory cells cycled circannually and were normal in number. Cytokine dysregulation was especially noted in one patient with systemic symptoms. Over a 2-year follow-up, cognitive deficits were observed in all patients following TGN1412 infusion. Some also had signs or symptoms of psychological, mucosal or immune dysregulation. These observations may discern immunopathology, treatment targets, and long-term monitoring strategies for other patients undergoing immunotherapy or with cytokine storm.- Published
- 2021
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15. A biorefinery for conversion of citrus peel waste into essential oils, pectin, fertilizer and succinic acid via different fermentation strategies.
- Author
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Patsalou M, Chrysargyris A, Tzortzakis N, and Koutinas M
- Subjects
- Actinobacillus, Fermentation, Fertilizers, Pectins, Succinic Acid, Citrus, Oils, Volatile
- Abstract
A process for the valorization of citrus peel waste (CPW) has been developed aiming to produce succinic acid and a series of added-value products through the biorefinery platform. CPW was subject to physicochemical and biological treatment to isolate essential oils (0.43%) and pectin (30.53%) as extractable products, pretreating the material for subsequent production of succinic acid that enabled application of remaining biorefinery residues (BR) as fertilizer substitute. Cellulose, hemicellulose and lignin contents of CPW accounted for 22.45%, 8.05% and 0.66% respectively, while acid hydrolysis reduced hemicellulose by 3.42% in BR. Moreover, essential oils extracted from CPW included 17 compounds, among which D-limonene reached 96.7%. The hydrolyzate generated was fermented for succinic acid production using Actinobacillus succinogenes. Different batch experiments demonstrated that the combined use of corn steep liquor (CSL) and vitamins in a lab-scale bioreactor resulted in product concentration and yield that reached 18.5 g L
-1 and 0.62 g g-1 respectively. Although simultaneous saccharification and fermentation (SSF) could not enhance succinic acid production, a fed-batch fermentation strategy increased succinic acid concentration and yield generating 22.4 g L-1 and 0.73 g g-1 respectively, while the mass of the platform chemical formed was enhanced by 27% as compared to the batch process. BR was explored as fertilizer substitute aiming to close the loop in the management of CPW towards development of a zero-waste process demonstrating that although the material imposed stress on plant growth, the content of potassium, phosphorus and nitrogen in the mixture increased., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)- Published
- 2020
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16. Resolution of alkaloid racemate: a novel microbial approach for the production of enantiopure lupanine via industrial wastewater valorization.
- Author
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Parmaki S, Tsipa A, Vasquez MI, Gonçalves JMJ, Hadjiadamou I, Ferreira FC, Afonso CAM, Drouza C, and Koutinas M
- Subjects
- Food Industry, Lupinus chemistry, Sparteine metabolism, Stereoisomerism, Wastewater chemistry, Biotransformation, Pseudomonas putida metabolism, Rhodococcus metabolism, Sparteine analogs & derivatives, Wastewater microbiology
- Abstract
Background: Lupanine is a plant toxin contained in the wastewater of lupine bean processing industries, which could be used for semi-synthesis of various novel high added-value compounds. This paper introduces an environmental friendly process for microbial production of enantiopure lupanine., Results: Previously isolated P. putida LPK411, R. rhodochrous LPK211 and Rhodococcus sp. LPK311, holding the capacity to utilize lupanine as single carbon source, were employed as biocatalysts for resolution of racemic lupanine. All strains achieved high enantiomeric excess (ee) of L-(-)-lupanine (> 95%), while with the use of LPK411 53% of the initial racemate content was not removed. LPK411 fed with lupanine enantiomers as single substrates achieved 92% of D-(+)-lupanine biodegradation, whereas L-(-)-lupanine was not metabolized. Monitoring the transcriptional kinetics of the luh gene in cultures supplemented with the racemate as well as each of the enantiomers supported the enantioselectivity of LPK411 for D-(+)-lupanine biotransformation, while (trans)-6-oxooctahydro-1H-quinolizine-3-carboxylic acid was detected as final biodegradation product from D-(+)-lupanine use. Ecotoxicological assessment demonstrated that lupanine enantiomers were less toxic to A. fischeri compared to the racemate exhibiting synergistic interaction., Conclusions: The biological chiral separation process of lupanine presented here constitutes an eco-friendly and low-cost alternative to widely used chemical methods for chiral separation.
- Published
- 2020
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17. A Citrus Peel Waste Biorefinery for Ethanol and Methane Production.
- Author
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Patsalou M, Samanides CG, Protopapa E, Stavrinou S, Vyrides I, and Koutinas M
- Subjects
- Anaerobiosis, Fatty Acids metabolism, Fermentation, Sugars analysis, Volatilization, Yeasts metabolism, Biotechnology methods, Citrus chemistry, Ethanol metabolism, Methane biosynthesis, Waste Products
- Abstract
This paper deals with the development of a citrus peel waste (CPW) biorefinery that employs low environmental impact technologies for production of ethanol and methane. Three major yeasts were compared for ethanol production in batch fermentations using CPW pretreated through acid hydrolysis and a combination of acid and enzyme hydrolysis. The most efficient conditions for production of CPW-based hydrolyzates included processing at 116 °C for 10 min. Pichia kudriavzevii KVMP10 achieved the highest ethanol production that reached 30.7 g L
-1 in fermentations conducted at elevated temperatures (42 °C). A zero-waste biorefinery was introduced by using solid biorefinery residues in repeated batch anaerobic digestion fermentations achieving methane formation of 342 mL gVS -1 (volatile solids). Methane production applying untreated and dried CPW reached a similar level (339-356 mL gVS -1 ) to the use of the side stream, demonstrating that the developed bioprocess constitutes an advanced alternative to energy intensive methods for biofuel production.- Published
- 2019
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18. Biodegradation and toxicity of emerging contaminants: Isolation of an exopolysaccharide-producing Sphingomonas sp. for ionic liquids bioremediation.
- Author
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Koutinas M, Vasquez MI, Nicolaou E, Pashali P, Kyriakou E, Loizou E, Papadaki A, Koutinas AA, and Vyrides I
- Subjects
- Aliivibrio fischeri drug effects, Animals, Biodegradation, Environmental, Chlorophyceae drug effects, Daphnia drug effects, Polysaccharides, Bacterial metabolism, RNA, Ribosomal, 16S genetics, Sphingomonas genetics, Sphingomonas isolation & purification, Environmental Pollutants metabolism, Environmental Pollutants toxicity, Ionic Liquids metabolism, Ionic Liquids toxicity, Sphingomonas metabolism
- Abstract
Ionic liquids (ILs) have been characterized as contaminants of emerging concern (CEC) that often resist biodegradation and impose toxicity upon environmental release. Sphingomonas sp. MKIV has been isolated as an extreme microorganism capable for biodegradation of major classes of ILs. Six imidazolium-, pyridinium- and ammonium-based ILs (pyridinium trifluoromethanesulfonate [Py][CF
3 SO3 ], 1-(4-pyridyl)pyridinium chloride [1-4PPy][Cl], 1-butyl-3-methylimidazolium bromide [BMIM][Br], 1-butyl-3-methylimidazolium methanesulfonate [BMIM][MeSO4 ], tetrabutylammonium iodide [n-Bu4 N][I] and tetrabutylammonium hexafluorophosphate [n-Bu4 N][PF6 ]) were used for microbial growth. The strain achieved 91% and 87% removal efficiency for cultures supplemented with 100 mg L-1 of [BMIM][MeSO4 ] and [n-Bu4 N][I] respectively. The metabolic activity of MKIV was inhibited following preliminary stages of cultures conducted using [BMIM][MeSO4 ], [BMIM][Br], [Py][CF3 SO3 ] and [n-Bu4 N][PF6 ], indicating potential accumulation of inhibitory metabolites. Thus, a comprehensive toxicological study of the six ILs on Aliivibrio fischeri, Daphnia magna and Raphidocelis subcapitata was conducted demonstrating that the compounds impose moderate and low toxicity. The end-products from [BMIM][MeSO4 ] and [n-Bu4 N][I] biodegradation were assessed using Aliivibrio fischeri, exhibiting increased environmental impact of the latter following biotreatment. MKIV produced 19.29 g L-1 of biopolymer, comprising mainly glucose and galacturonic acid, from 25 g L-1 of glucose indicating high industrial significance for bioremediation and exopolysaccharide production., (Copyright © 2018 Elsevier B.V. All rights reserved.)- Published
- 2019
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19. Optimal bioprocess design through a gene regulatory network - Growth kinetic hybrid model: Towards replacing Monod kinetics.
- Author
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Tsipa A, Koutinas M, Usaku C, and Mantalaris A
- Subjects
- Gene Regulatory Networks, Genes, Bacterial, Metabolic Engineering methods, Models, Genetic, Pseudomonas putida genetics, Pseudomonas putida metabolism
- Abstract
Currently, design and optimisation of biotechnological bioprocesses is performed either through exhaustive experimentation and/or with the use of empirical, unstructured growth kinetics models. Whereas, elaborate systems biology approaches have been recently explored, mixed-substrate utilisation is predominantly ignored despite its significance in enhancing bioprocess performance. Herein, bioprocess optimisation for an industrially-relevant bioremediation process involving a mixture of highly toxic substrates, m-xylene and toluene, was achieved through application of a novel experimental-modelling gene regulatory network - growth kinetic (GRN-GK) hybrid framework. The GRN model described the TOL and ortho-cleavage pathways in Pseudomonas putida mt-2 and captured the transcriptional kinetics expression patterns of the promoters. The GRN model informed the formulation of the growth kinetics model replacing the empirical and unstructured Monod kinetics. The GRN-GK framework's predictive capability and potential as a systematic optimal bioprocess design tool, was demonstrated by effectively predicting bioprocess performance, which was in agreement with experimental values, when compared to four commonly used models that deviated significantly from the experimental values. Significantly, a fed-batch biodegradation process was designed and optimised through the model-based control of TOL Pr promoter expression resulting in 61% and 60% enhanced pollutant removal and biomass formation, respectively, compared to the batch process. This provides strong evidence of model-based bioprocess optimisation at the gene level, rendering the GRN-GK framework as a novel and applicable approach to optimal bioprocess design. Finally, model analysis using global sensitivity analysis (GSA) suggests an alternative, systematic approach for model-driven strain modification for synthetic biology and metabolic engineering applications., (Copyright © 2018. Published by Elsevier Inc.)
- Published
- 2018
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20. Bioconversion of alkaloids to high-value chemicals: Comparative analysis of newly isolated lupanine degrading strains.
- Author
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Parmaki S, Vyrides I, Vasquez MI, Hartman V, Zacharia I, Hadjiadamou I, Barbeitos CBM, Ferreira FC, Afonso CAM, Drouza C, and Koutinas M
- Subjects
- Aliivibrio fischeri metabolism, Alkaloids analysis, Alkaloids chemistry, Animals, Daphnia metabolism, Magnoliopsida metabolism, Pseudomonas putida metabolism, Sparteine analysis, Sparteine chemistry, Sparteine metabolism, Alkaloids metabolism, Biodegradation, Environmental, Sparteine analogs & derivatives
- Abstract
This work explores the potential for development of a lupanine valorization process evaluating different isolated microorganisms for their capacity to metabolize the alkaloid. Ecotoxicological assessment demonstrated that lupanine is toxic for Vibrio fischeri and Daphnia magna exhibiting EC
50 values of 89 mg L-1 and 47 mg L-1 respectively, while acting both as growth inhibitor for a monocotyledonous and as promoter for a dicotyledonous plant. Among the eight aerobic and anaerobic strains isolated and identified Rhodococcus rhodochrous LPK211 achieved 81% removal for 1.5 g L-1 lupanine, while no end-products were detected by NMR constituting a promising microorganism for lupanine biodegradation. Moreover, Rhodococcus ruber LPK111 and Rhodococcus sp. LPK311 exhibited 66% and 71% of removal respectively, including potential formation of lupanine N-oxide. Pseudomonas putida LPK411 reached 80% of lupanine removal and generated three fermentation products potentially comprising 17-oxolupanine and lupanine derivatives with open ring structures enabling the development of alkaloid valorization processes., (Copyright © 2017 Elsevier Ltd. All rights reserved.)- Published
- 2018
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21. Application of commercial and non-commercial immobilized lipases for biocatalytic production of ethyl lactate in organic solvents.
- Author
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Koutinas M, Yiangou C, Osório NM, Ioannou K, Canet A, Valero F, and Ferreira-Dias S
- Subjects
- Biocatalysis, Enzymes, Immobilized, Esterification, Solvents, Lactates, Lipase
- Abstract
This study explores the potential for enhancing the production of ethyl lactate (EL), a green solvent, through enzymatic esterification. Different solvents were compared as organic media for conversion of lactate and ethanol into EL, catalyzed by Novozym 435. Chloroform and hexane were the most effective in low acid concentrations (0.01-0.1M) exhibiting maximum EL yields of 88% and 75% respectively. The yield of EL improved as the solvent's LogP increased up to a value of 2. Non-commercial immobilized biocatalysts consisting heterologous Rhizopous oryzae (rROL) and Candida rugosa (CRL) lipases immobilized on hydrophobic supports were compared to commercial biocatalysts clarifying that Novozym 435 and Lipozyme RM IM could be efficiently applied. Operational stability tests were conducted using Novozym 435, which retained higher activity in chloroform as compared to hexane. Although non-commercial biocatalysts were not competitive in esterification, they exhibited significant activity towards hydrolysis constituting a valuable alternative to higher-cost options., (Copyright © 2017.)
- Published
- 2018
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22. The impact of succinate trace on pWW0 and ortho-cleavage pathway transcription in Pseudomonas putida mt-2 during toluene biodegradation.
- Author
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Tsipa A, Koutinas M, Vernardis SI, and Mantalaris A
- Subjects
- Batch Cell Culture Techniques, Biodegradation, Environmental, Bioreactors, Gene Expression Regulation, Bacterial, Kinetics, Metabolic Networks and Pathways, Promoter Regions, Genetic, Pseudomonas putida genetics, Transcription, Genetic, Environmental Pollutants metabolism, Plasmids genetics, Pseudomonas putida metabolism, Succinates metabolism, Toluene metabolism
- Abstract
Toluene is a pollutant catabolised through the interconnected pWW0 (TOL) and ortho-cleavage pathways of Pseudomonas putida mt-2, while upon succinate and toluene mixtures introduction in batch cultures grown on M9 medium, succinate was previously reported as non-repressing. The effect of a 40 times lower succinate concentration, as compared to literature values, was explored through systematic real-time qPCR monitoring of transcriptional kinetics of the key TOL Pu, Pm and ortho-cleavage PbenR, PbenA promoters in mixed-substrate experiments. Even succinate trace inhibited transcription leading to bi-modal promoters expression. Potential carbon catabolite repression mechanisms and novel expression patterns of promoters were unfolded. Lag phase was shortened and biomass growth levels increased compared to sole toluene biodegradation suggesting enhanced pollutant removal efficiency. The study stressed the noticeable effect of a preferred compound's left-over on the main route of a bioprocess, revealing the beneficiary supply of low preferred substrates concentrations to design optimal bioremediation strategies., (Copyright © 2017 Elsevier Ltd. All rights reserved.)
- Published
- 2017
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23. Transcriptional kinetics of the cross-talk between the ortho-cleavage and TOL pathways of toluene biodegradation in Pseudomonas putida mt-2.
- Author
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Tsipa A, Koutinas M, Pistikopoulos EN, and Mantalaris A
- Subjects
- Kinetics, Metabolic Networks and Pathways genetics, Promoter Regions, Genetic genetics, Pseudomonas putida metabolism, Signal Transduction genetics, Gene Expression Regulation, Bacterial genetics, Plasmids genetics, Pseudomonas putida genetics, Toluene metabolism, Transcription, Genetic genetics
- Abstract
The TOL plasmid promoters are activated by toluene leading to gene expression responsible for the degradation of the environmental signal. Benzoate is formed as an intermediate, activating the BenR protein of the chromosomal ortho-cleavage pathway that up-regulates the chromosomal PbenA promoter and the TOL Pm promoter resulting in cross-talk between the two networks. Herein, the transcriptional kinetics of the PbenR and PbenA promoters in conjunction with TOL promoters was monitored by real-time PCR during toluene biodegradation of different concentrations in batch cultures. The cross-talk between the two pathways was indicated by the simultaneous maximal expression of the Pm and PbenR promoters, as well as the transcriptional activation from PbenA occurring prior to PbenR, which indicates the potential up-regulation of PbenA by the TOL XylS protein. The repressory effect of toluene on Pr was evident for concentrations higher than 0.3mM suggesting a threshold value for restoring the promoter's activity, while all the other promoters followed a specific expression pattern, regardless of the initial inducer concentration. Induction of the system with higher toluene concentrations revealed an oscillatory behaviour of Pm, the expression of which remained at high levels until the late exponential phase, demonstrating a novel function of this network., (Copyright © 2016. Published by Elsevier B.V.)
- Published
- 2016
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24. Novel Halomonas sp. B15 isolated from Larnaca Salt Lake in Cyprus that generates vanillin and vanillic acid from ferulic acid.
- Author
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Vyrides I, Agathangelou M, Dimitriou R, Souroullas K, Salamex A, Ioannou A, and Koutinas M
- Subjects
- Cyprus, Halomonas classification, Halomonas genetics, Lakes analysis, Molecular Sequence Data, Phylogeny, Sodium Chloride analysis, Sodium Chloride metabolism, Benzaldehydes metabolism, Coumaric Acids metabolism, Halomonas isolation & purification, Halomonas metabolism, Lakes microbiology, Vanillic Acid metabolism
- Abstract
Vanillin is a high value added product with many applications in the food, fragrance and pharmaceutical industries. A natural and low-cost method to produce vanillin is by microbial bioconversions through ferulic acid. Until now, limited microorganisms have been found capable of bioconverting ferulic acid to vanillin at high yield. This study aimed to screen halotolerant strains of bacteria from Larnaca Salt Lake which generate vanillin and vanillic acid from ferulic acid. From a total of 50 halotolenant/halophilic strains 8 grew in 1 g/L ferulic acid and only 1 Halomonas sp. B15 and 3 Halomonas elognata strains were capable of bioconverting ferulic acid to vanillic acid at 100 g NaCl/L. The highest vanillic acid (365 mg/L) at these conditions generated by Halomonas sp. B15 which corresponds to ferulic acid bioconversion yield of 36.5%. Using the resting cell technique with an initial ferulic acid concentration of 0.5 g/L at low salinity, the highest production of vanillin (245 mg/L) took place after 48 h, corresponding to a bioconversion yield of 49%. This is the first reported Halomonas sp. with high yield of vanillin production from ferulic acid at low salinity.
- Published
- 2015
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25. Development of a hybrid fermentation-enzymatic bioprocess for the production of ethyl lactate from dairy waste.
- Author
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Koutinas M, Menelaou M, and Nicolaou EN
- Subjects
- Biofuels, Enzymes, Immobilized, Esterification, Ethanol metabolism, Feasibility Studies, Fungal Proteins, Kluyveromyces metabolism, Lactobacillus metabolism, Liquid-Liquid Extraction, Solutions, Solvents, Temperature, Toluene chemistry, Water chemistry, Biotechnology methods, Dairying, Fermentation, Lactates metabolism, Lipase metabolism, Waste Products
- Abstract
This work explores the potential for the development of a hybrid fermentation-enzymatic process for the production of ethyl lactate from dairy waste. Cheese whey was used in Kluyveromyces marxianus and Lactobacillus bulgaricus batch cultures to produce ethanol and lactic acid respectively. Subsequently, the fermentation products were transferred into an organic phase through liquid-liquid extraction and ethyl lactate was formed in an esterification reaction catalyzed by lipases. The production of ethanol and lactic acid achieved under different conditions was 23gL(-1) and 29gL(-1), respectively. Furthermore, the efficiency of various organic solvents for the esterification reaction was evaluated and toluene was chosen for application in the process. The effect of water content was determined aiming to maximize the product yield and 40mgml(-1) was the optimal enzyme concentration. The bioprocess achieved maximum conversion of 33% constituting a valuable alternative to the application of energy demanding chemically derived methods., (Copyright © 2014 Elsevier Ltd. All rights reserved.)
- Published
- 2014
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26. Bioprocess systems engineering: transferring traditional process engineering principles to industrial biotechnology.
- Author
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Koutinas M, Kiparissides A, Pistikopoulos EN, and Mantalaris A
- Abstract
The complexity of the regulatory network and the interactions that occur in the intracellular environment of microorganisms highlight the importance in developing tractable mechanistic models of cellular functions and systematic approaches for modelling biological systems. To this end, the existing process systems engineering approaches can serve as a vehicle for understanding, integrating and designing biological systems and processes. Here, we review the application of a holistic approach for the development of mathematical models of biological systems, from the initial conception of the model to its final application in model-based control and optimisation. We also discuss the use of mechanistic models that account for gene regulation, in an attempt to advance the empirical expressions traditionally used to describe micro-organism growth kinetics, and we highlight current and future challenges in mathematical biology. The modelling research framework discussed herein could prove beneficial for the design of optimal bioprocesses, employing rational and feasible approaches towards the efficient production of chemicals and pharmaceuticals.
- Published
- 2013
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27. Linking genes to microbial growth kinetics: an integrated biochemical systems engineering approach.
- Author
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Koutinas M, Kiparissides A, Silva-Rocha R, Lam MC, Martins Dos Santos VA, de Lorenzo V, Pistikopoulos EN, and Mantalaris A
- Subjects
- Gene Expression Regulation, Bacterial genetics, Gene Expression Regulation, Enzymologic genetics, Kinetics, Oxidation-Reduction, Plasmids genetics, Plasmids metabolism, Transcription, Genetic genetics, Xylenes metabolism, Models, Biological, Pseudomonas putida enzymology, Pseudomonas putida genetics, Pseudomonas putida growth & development
- Abstract
The majority of models describing the kinetic properties of a microorganism for a given substrate are unstructured and empirical. They are formulated in this manner so that the complex mechanism of cell growth is simplified. Herein, a novel approach for modelling microbial growth kinetics is proposed, linking biomass growth and substrate consumption rates to the gene regulatory programmes that control these processes. A dynamic model of the TOL (pWW0) plasmid of Pseudomonas putida mt-2 has been developed, describing the molecular interactions that lead to the transcription of the upper and meta operons, known to produce the enzymes for the oxidative catabolism of m-xylene. The genetic circuit model was combined with a growth kinetic model decoupling biomass growth and substrate consumption rates, which are expressed as independent functions of the rate-limiting enzymes produced by the operons. Estimation of model parameters and validation of the model's predictive capability were successfully performed in batch cultures of mt-2 fed with different concentrations of m-xylene, as confirmed by relative mRNA concentration measurements of the promoters encoded in TOL. The growth formation and substrate utilisation patterns could not be accurately described by traditional Monod-type models for a wide range of conditions, demonstrating the critical importance of gene regulation for the development of advanced models closely predicting complex bioprocesses. In contrast, the proposed strategy, which utilises quantitative information pertaining to upstream molecular events that control the production of rate-limiting enzymes, predicts the catabolism of a substrate and biomass formation and could be of central importance for the design of optimal bioprocesses., (Copyright © 2011 Elsevier Inc. All rights reserved.)
- Published
- 2011
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28. Modelling the Delta1/Notch1 pathway: in search of the mediator(s) of neural stem cell differentiation.
- Author
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Kiparissides A, Koutinas M, Moss T, Newman J, Pistikopoulos EN, and Mantalaris A
- Subjects
- Basic Helix-Loop-Helix Transcription Factors chemistry, Basic Helix-Loop-Helix Transcription Factors genetics, Basic Helix-Loop-Helix Transcription Factors metabolism, Computational Biology, Protein Multimerization, Protein Structure, Quaternary, RNA, Messenger genetics, RNA, Messenger metabolism, Cell Differentiation, Intracellular Signaling Peptides and Proteins metabolism, Membrane Proteins metabolism, Models, Biological, Neural Stem Cells cytology, Neural Stem Cells metabolism, Receptor, Notch1 metabolism, Signal Transduction
- Abstract
The Notch1 signalling pathway has been shown to control neural stem cell fate through lateral inhibition of mash1, a key promoter of neuronal differentiation. Interaction between the Delta1 ligand of a differentiating cell and the Notch1 protein of a neighbouring cell results in cleavage of the trans-membrane protein, releasing the intracellular domain (NICD) leading to the up regulation of hes1. Hes1 homodimerisation leads to down regulation of mash1. Most mathematical models currently represent this pathway up to the formation of the HES1 dimer. Herein, we present a detailed model ranging from the cleavage of the NICD and how this signal propagates through the Delta1/Notch1 pathway to repress the expression of the proneural genes. Consistent with the current literature, we assume that cells at the self renewal state are represented by a stable limit cycle and through in silico experimentation we conclude that a drastic change in the main pathway is required in order for the transition from self-renewal to differentiation to take place. Specifically, a model analysis based approach is utilised in order to generate hypotheses regarding potential mediators of this change. Through this process of model based hypotheses generation and testing, the degradation rates of Hes1 and Mash1 mRNA and the dissociation constant of Mash1-E47 heterodimers are identified as the most potent mediators of the transition towards neural differentiation.
- Published
- 2011
- Full Text
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29. The regulatory logic of m-xylene biodegradation by Pseudomonas putida mt-2 exposed by dynamic modelling of the principal node Ps/Pr of the TOL plasmid.
- Author
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Koutinas M, Lam MC, Kiparissides A, Silva-Rocha R, Godinho M, Livingston AG, Pistikopoulos EN, de Lorenzo V, Dos Santos VA, and Mantalaris A
- Subjects
- Biodegradation, Environmental, Models, Theoretical, Molecular Structure, Transcription, Genetic, Xylenes chemistry, Gene Expression Regulation, Bacterial, Models, Biological, Plasmids genetics, Plasmids metabolism, Pseudomonas putida genetics, Pseudomonas putida metabolism, Xylenes metabolism
- Abstract
The structure of the extant transcriptional control network of the TOL plasmid pWW0 born by Pseudomonas putida mt-2 for biodegradation of m-xylene is far more complex than one would consider necessary from a mere engineering point of view. In order to penetrate the underlying logic of such a network, which controls a major environmental cleanup bioprocess, we have developed a dynamic model of the key regulatory node formed by the Ps/Pr promoters of pWW0, where the clustering of control elements is maximal. The model layout was validated with batch cultures estimating parameter values and its predictive capability was confirmed with independent sets of experimental data. The model revealed how regulatory outputs originated in the divergent and overlapping Ps/Pr segment, which expresses the transcription factors XylS and XylR respectively, are computed into distinct instructions to the upper and lower catabolic xyl operons for either simultaneous or stepwise consumption of m-xylene and/or succinate. In this respect, the model reveals that the architecture of the Ps/Pr is poised to discriminate the abundance of alternative and competing C sources, in particular m-xylene versus succinate. The proposed framework provides a first systemic understanding of the causality and connectivity of the regulatory elements that shape this exemplary regulatory network, facilitating the use of model analysis towards genetic circuit optimization.
- Published
- 2010
- Full Text
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30. The use of an oil absorber as a strategy to overcome starvation periods in degrading 1,2-dichloroethane in waste gas.
- Author
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Koutinas M, Baptista II, Peeva LG, Ferreira Jorge RM, and Livingston AG
- Subjects
- Absorption, Air Pollutants chemistry, Biodegradation, Environmental, Bioreactors, Ethylene Dichlorides chemistry, Industrial Waste, Models, Theoretical, Plant Oils metabolism, Xanthobacter metabolism, Air Pollutants metabolism, Ethylene Dichlorides metabolism, Plant Oils chemistry
- Abstract
This work investigates the use of an oil absorber as an operational strategy in vapor phase bioreactors exposed to starvation periods, during the treatment of inhibitory pollutants. After being exposed to 1,2-dichloroethane (DCE) starvation periods, the response and stability of a combined oil-absorber-bioscrubber (OAB) system was compared to that of a bioscrubber only (BO) system. In the BO system, after a 5.2 days starvation period, the DCE removal efficiency was reduced to 12%, and 6 days were needed to recover the initial removal efficiency when the DCE feed resumed. The total organic discharged (TOD(DCE)) was 16,500 g(DCE) m(bioscrubber) (-3) after the DCE starvation. Biomass analysis performed using fluorescence in situ hybridisation (FISH) showed that the microbial activity was significantly reduced during the starvation period and that 5 days were needed to recover the initial activity, after the re-introduction of DCE. In contrast, the performance of the OAB system was stable during 5.2 days of DCE starvation. The DCE removal efficiency was not affected when the DCE feed resumed and the TOD(DCE) was significantly reduced to 2,850 g(DCE) m(bioscrubber) (-3). During starvation, the activity of the microbial culture in the OAB system showed a substantially lower decrease than in the BO system and recovered almost immediately the initial activity after the re-introduction of DCE. Additionally, a mathematical model describing the performance of the OAB system was developed. The results of this study show that the OAB system can effectively sustain the biological treatment of waste gas during starvation periods of inhibitory pollutants.
- Published
- 2007
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31. An oil-absorber-bioscrubber system to stabilize biotreatment of pollutants present in waste gas. Fluctuating loads of 1,2-dichloroethane.
- Author
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Koutinas M, Martin J, Peeva LG, Mantalaris A, and Livingston AG
- Subjects
- Air Pollutants metabolism, Ethylene Dichlorides metabolism, Air Pollutants chemistry, Ethylene Dichlorides chemistry, Oils, Xanthobacter metabolism
- Abstract
Biotreatment technologies offer a cost-effective and efficient method for dealing with point-source releases of solvents. However, a major problem hampering these technologies is the fluctuating pollutant loads, which is especially critical for inhibitory pollutants. Provision of biotreatment systems able to cope with this problem is a significant technological and environmental challenge. This study investigates the potential for an absorber to act as buffer for shock loadings of inhibitory pollutants in waste-gas streams undergoing biological treatment. 1,2-Dichloroethane (DCE) was used as an example of a toxic and inhibitory organic pollutant. The stability of a combined oil-absorber-bioscrubber (OAB) system was compared to that of a bioscrubber only (BO) system when each was subjected to shock loads of DCE. The BO system was inoculated with Xanthobacter autotrophicus strain GJ10 and was submitted to sharp, sequential pulses in DCE inlet load, which caused system instability. Complete inhibition of the BO process occurred for a 3 h DCE pulse, leading to 9125 g of DCE m(-3)bioscrubber total organic discharged (TODDCE). Following the pulse, fluorescence in situ hybridization (FISH) showed that the active strain GJ10 was effectively washed-out. In contrast, the performance of the OAB system was stable during DCE shock loads. The carbon dioxide production remained stable, and low levels of effluent DCE and total organic carbon concentrations were found. For the 3 h pulse TODDCE was only 173 g of DCE m(-3)bioscrubber, and FISH indicated that the GJ10 strain remained active. We conclude that the OAB system offers an effective solution to the biological treatment of waste-gas containing fluctuating pollutant concentrations.
- Published
- 2006
- Full Text
- View/download PDF
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