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Research Project
BIOLOGICAL PRODUCTION OF INDUSTRIALLY RELEVANT FLUOROORGANIC COMPOUNDS
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Publications
Bioaugmentation of a rotating biological contactor for degradation of 2-fluorophenol
Publication . Duque, Anouk F.; Bessa, Vânia S.; Carvalho, Maria F.; Castro, Paula M. L.
The performance of a laboratory scale rotating biological contactor (RBC) towards shock loadings of 2-fluorophenol (2-FP) was investigated. During a period of ca. 2 months organic shock loadings of 25 mg L-1 of 2-FP were applied to the RBC. As no biodegradation of 2-FP was observed, bioaugmentation of the RBC with a 2-FP degrading strain was carried out and, along ca. 6 months, organic shock loadings within a range of 25-200 mg L-1 of 2-FP were applied. Complete biodegradation of 50 mg L-1 of 2-FP was observed during operation of the reactor. The RBC showed to be robust towards starvation periods, as after ca. 1 month of non-supply of the target compound, the reactor resumed 2-FP degradation. The inoculated strain was retained within the biofilm in the disks, as the 2-FP degrading strain was recovered from the biofilm by the end of the experiment, thus bioaugmentation was successfully achieved.
Co-metabolic degradation of chlorobenzene by the fluorobenzene degrading wild strain Labrys portucalensis
Publication . Moreira, Irina S.; Amorim, Catarina L.; Carvalho, Maria F.; Castro, Paula M.L.
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 mmol 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 mmol 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 mmol l 1 day 1 and 126 16.2 mmol 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.
Isolation and characterization of a Rhodococcus strain able to degrade 2-fluorophenol
Publication . Duque, Anouk F.; Hasan, Syed A.; Bessa, Vânia S.; Carvalho, Maria F.; Samin, Ghufrana; Janssen, Dick B.; Castro, Paula M. L.
A pure bacterial culture able to utilize 2- fluorophenol (2-FP) as sole carbon and energy source was isolated by selective enrichment from sediments collected from a contaminated site in Northern Portugal. 16S rRNA gene analysis showed that the organism (strain FP1) belongs to the genus Rhodococcus. When grown aerobically on 2-FP, growth kinetics of strain FP1 followed the Luong model. An inhibitory effect of increasing 2-FP concentrations was observed with no growth occurring at 2- FP levels higher than ca. 4 mM. Rhodococcus strain FP1 was able to degrade a range of other organofluorine compounds, including 2-fluorobenzoate, 3-fluorobenzoate, 4-fluorobenzoate, 3-fluorophenol, 4-fluorophenol, 3-fluorocatechol, and 4-fluorocatechol, as well as chlorinated compounds such as 2-chlorophenol and 4-chlorophenol. Experiments with cell-free extracts and partially purified enzymes indicated that the first step of 2-fluorophenol metabolism was conversion to 3-fluorocatechol, suggesting an unusual pathway for fluoroaromatic metabolism. To our knowledge, this is the first time that utilization of 2-FP as a growth substrate by a pure bacterial culture is reported.
Effect of the metals iron, copper and silver on fluorobenzene biodegradation by Labrys portucalensis
Publication . Moreira, Irina S.; Amorim, Catarina L.; Carvalho, Maria F.; Ferreira, António C.; Afonso, Carlos M.; Castro, Paula M. L.
Organic and metallic pollutants are ubiquitous in the environment. Many metals are reported to be toxic to microorganisms and to inhibit biodegradation. The effect of the metals iron, copper and silver on the metabolism of Labrys portucalensis F11 and on fluorobenzene (FB) biodegradation was examined. The results indicate that the addition of 1 mM of Fe2+ to the culture medium has a positive effect on bacterial growth and has no impact in the biodegradation of 1 and 2 mM of FB. The presence of 1 mM of Cu2+ was found to strongly inhibit the growth of F11 cultures and to reduce the biodegradation of 1 and 2 mM of FB to ca. 50 %, with 80 % of stoichiometrically expected fluoride released. In the experiments with resting cells, the FB degraded (from 2 mM supplied) was reduced ca. 20 % whereas the fluoride released was reduced to 45 % of that stoichiometrically expected. Ag+ was the most potent inhibitor of FB degradation. In experiments with growing cells, the addition of 1 mM of Ag+ to the culture medium containing 1 and 2 mM of FB resulted in no fluoride release, whereas FB degradation was only one third of that observed in control cultures. In the experiments with resting cells, the addition of Ag+ resulted in 25 % reduction in substrate degradation and fluoride release was only 20 % of that stoichiometrically expected. The accumulation of catechol and 4-fluorocatechol in cultures supplemented with Cu2+ or Ag+ suggest inhibition of the key enzyme of FB metabolism-catechol 1,2-dioxygenase.
Bacterial degradation of moxifloxacin in the presence of acetate as a bulk substrate
Publication . Carvalho, M. F.; Maia, A. S.; Tiritan, M. E.; Castro, P. M. L.
Fluoroquinolones constitute a group of emerging pollutants and their occurrence in different environmental
compartments is becoming object of increasing public concern due to their ecotoxicological effects
and the potential to develop resistant bacteria. This study aimed to investigate the biodegradation
of moxifloxacin (MOX), for which studies in the literature are very scarce. An activated sludge (AS)
consortium and three bacterial strains able to degrade fluoroaromatic compounds e strains F11, FP1 and
S2 e were tested. Biodegradation studies were conducted using acetate as a bulk carbon source. Strain
F11 showed the highest biodegradation capacity, being able to completely consume and dehalogenate
7.5 mM of the target antibiotic when daily co-supplemented with acetate present as a readily degradable
organic substrate in wastewaters. MOX could be used by strain F11 as a sole nitrogen source but the
presence of an external nitrogen source in the culture medium was essential for complete biodegradation.
Strain F11 was capable of completely consuming MOX in a range between 2 and 11 mM, although
stoichiometric fluoride release was not obtained for the highest tested concentration. The antibacterial
activity of residual MOX and of the metabolic products potentially resultant from the biodegradation
process was investigated by agar diffusion tests, demonstrating that MOX biodegradation is associated
with the elimination of the antibacterial properties of the target antibiotic and of the produced metabolites,
which is an important result, as the activity of antibiotics and/or their metabolites in the
environment, even at low levels, may lead to the development of resistant bacterial strains. Overall, the
results obtained in this study suggest that strain F11 is a promising microorganism for the treatment of
waters contaminated with MOX, where it could be used for bioaugmentation/bioremediation purposes.
To the best of our knowledge, this is the first study reporting complete removal and dehalogenation of
MOX by a single microorganism.
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Funding agency
Fundação para a Ciência e a Tecnologia
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Funding Award Number
SFRH/BPD/44670/2008