• Journal of Microbiology
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• v.34 no.4
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• pp.363-369
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• 1996

As basic study to evaluate the treatability of oil-contaminated environment with bacteria, isolation and characterization of crude oil-degrading bacterium were carried out. A bacterial strain SH-14 capable of degrading crude oil was isolated from contaminated soils by enrichment culture technique and identified as Acinetobacter sp. by morphological, cultural and biochemical characteristics, and so named Acinetobacter sp. SH-14. The optimal medium composition and cultural conditions for the growth and emulsification of crude oil by Acinetobacter sp. SH-14 used were crude oil of 2.0%, $KNO_3$ of 0.2%, $K_2HPO_4$ of 0.05%, and $MgSO_4\;{\cdot}\;7H_2O$ of 1.0%, along with initial pH 7.0 at $30^{\circ}C$. Acinetobacter sp. SH-14 showed to be resistant to chloramphenicol and utilized various hydrocarbons such as dodecane, hexadecane, isooctane, cyclo-hexane etc., as a sole carbon source. Acinetobacter sp. SH-14 harbored a single plasmid. By agarose gel electrophoresis and curing experiment it was found that the genes for crude oil components degradation were encoded on the plasmid.

• Proceedings of the Korean Society of Soil and Groundwater Environment Conference
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• 2002.09a
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• pp.310-314
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• 2002

To investigate the effect of on-site bioremediation in soil that have been contaminated by hydrocarbon fuel spills, petroleum-degrading bacteria isolated from soil around petroleum chemical industry and microbial agents were constructed. We investigated biopiles for on-site bioremediation of soil contaminated (5000 mg per kg) with bunker A fuel in five independent lab-scale experiments. Five biopile units constituting the following treatments: (1) control with no nutrients and microbial agents (2) microbial agent M plus nutrients (3) microbial agent C plus nutrients (4) only microbial agent C (5) control with only nutrients. The results were highly different one another. After 30 days in treatments with optimal condition, total petroleum hydrocarbons were reduced to below 10 mg per kg of soil at the biopile units mixed with microbial agents, but control biopile units show that were reduced from 1,105 to 2,588 mg per kg of soil. Our results show that microbial agents at on-site bioremediation of fuel-contaminated soil is highly effective.

• Journal of Life Science
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• v.21 no.11
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• pp.1511-1517
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• 2011

We isolated a potent phenanthrene-degrading bacterium from oil-contaminated soils of Suzhou, China, and assessed the potential use of these bacteria for bioremediation of soils contaminated by polycyclic aromatic hydrocarbons (PAHs) in a microcosm. Based on 16S rDNA sequencing, we identified this bacteria as Sphigobacterium sp. SW-09. By PCR amplification, we also identified catechol 2,3-dioxygenase genes (nahH genes) mediating PAH degradation. Staphylococcus sp. KW-07, which has been identified in our previous study, showed potential for use in bioremediation of oil-contaminated soils. In this experiment, we compared the rate of phenanthrene-degradation between Staphylococcus sp. KW-07 and Sphingobacterium sp. SW-09 in a microcosm condition. Newly isolated Sphingobacterium sp. SW-09 showed a higher phenanthrene-degradation rate than that of Staphylococcus sp. KW-07 in soil microcosms. Together, our results suggest that the Sphingobacterim sp. SW-09 strain isolated from the Suzhou area may also be useful in bioremediation of PAH-contaminated soils.

• Journal of Microbiology and Biotechnology
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• v.22 no.3
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• pp.283-291
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• 2012

Bioremediation efforts often rely on the application of surfactants to enhance hydrocarbon bioavailability. However, synthetic surfactants can sometimes be toxic to degrading microorganisms, thus reducing the clearance rate of the pollutant. Therefore, surfactant-resistant bacteria can be an important tool for bioremediation efforts of hydrophobic pollutants, circumventing the toxicity of synthetic surfactants that often delay microbial bioremediation of these contaminants. In this study, we screened a natural surfactant-rich compartment, the estuarine surface microlayer (SML), for cultivable surfactant-resistant bacteria using selective cultures of sodium dodecyl sulfate (SDS) and cetyl trimethylammonium bromide (CTAB). Resistance to surfactants was evaluated by colony counts in solid media amended with critical micelle concentrations (CMC) of either surfactants, in comparison with non-amended controls. Selective cultures for surfactant-resistant bacteria were prepared in mineral medium also containing CMC concentrations of either CTAB or SDS. The surfactantresistant isolates obtained were tested by PCR for the Pseudomonas genus marker gacA gene and for the naphthalene-dioxygenase-encoding gene ndo. Isolates were also screened for biosurfactant production by the atomized oil assay. A high proportion of culturable bacterioneuston was tolerant to CMC concentrations of SDS or CTAB. The gacA-targeted PCR revealed that 64% of the isolates were Pseudomonads. Biosurfactant production in solid medium was detected in 9.4% of tested isolates, all affiliated with genus Pseudomonas. This study shows that the SML is a potential source of surfactant-resistant and biosurfactant-producing bacteria in which Pseudomonads emerge as a relevant group.

• Journal of Environmental Science International
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• v.8 no.4
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• pp.451-456
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• 1999

Microorganisms utilizing petroleum as substrate were screened from the seawater in Pusan coastal area. Among them, fifty strains utilized bunker-A oil as a sole carbon and energy source. Five of these fifty strains were selected to experiment this study. According to the taxonomic characteristics of its morphological, cultural and biochemical properties, the selected stains were named Pseudomonas sp. EL-12, Flavobacterium sp. EL-15, Acinetobacter sp. EL-18, Enterobacter sp. EL-27 and Micrococcus sp. EL-43, respectively. The optimal medium compositions and cultural conditions for assimilation of bunker-A oil by the selected strains were 1.5-2% bunker-A oil, 0.1% $NH_4NO_3$, 1-1.5% $MgSO_4$.$7H_2O$, 0.05-0.15% KCl, 0.1-0.15% $CaCl_2$.$2H_2O$, 2.5-3.5% NaCl, initial pH 8-9, temperature 3$0^{\circ}C$ and aeration, respectively. The utilization and degradation characteristics on the various hydrocarbons by the selected stains were showed that bunker oil, n-alkane and branched alkane compounds were highly activity than cyclic alkane and aromatic hydrocarbon compounds.

• Journal of Soil and Groundwater Environment
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• v.19 no.5
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• pp.9-17
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• 2014

This paper reported the use of real-time polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), and the culture-based method in the intrinsic bioremediation study at a petroleum contaminated site. The study showed that phenol hydroxylase gene was detected in groundwater contaminated with benzene, toluene, ethylbenzene, xylene isomers (BTEX) and methyl tert-butyl ether (MTBE). This indicated that intrinsic bioremediation occurred at the site. DGGE analyses revealed that the petroleum-hydrocarbon plume caused the variation in microbial communities. MTBE degraders including Pseudomonas sp. NKNU01, Bacillus sp. NKNU01, Klebsiella sp. NKNU01, Enterobacter sp. NKNU01, and Enterobacter sp. NKNU02 were isolated from the contaminated groundwater using the cultured-based method. Among these five strains, Enterobacter sp. NKNU02 is the most effective stain at degrading MTBE without the addition of pentane. The MTBE biodegradation experiment indicated that the isolated bacteria were affected by propane. Biodegradation of MTBE was decreased but not totally inhibited in the mixtures of BTEX. Enterobacter sp. NKNU02 degraded about 60% of MTBE in the bioreactor study. Tert-butyl alcohol (TBA), acetic acid, 2-propanol, and propenoic acid were detected using gas chromatography/mass spectrometry during MTBE degraded by the rest cells of Enterobacter sp. NKNU02. The effectiveness of bioremediation of MTBE was assessed for potential field-scale application.

• 한국생물공학회:학술대회논문집
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• 2000.04a
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• pp.89-91
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• 2000

Polynuclear aromatic hydrocarbon (PAH) compounds are highly carcinogenic chemicals and common groundwater contaminants that are observed to persist in soils. The adherence and slow release of PAHs in soil is an obstacle to remediation and complicates the assessment of cleanup standards and risks. Biological degradation of PAHs in soil has been an area of active research because biological treatment may be less costly than conventional pumping technologies or excavation and thermal treatment. Biological degradation also offers the advantage to transform PAHs into non-toxic products such as biomass and carbon dioxide. Ample evidence exists for aerobic biodegradation of PAHs and many bacteria capable of degrading PAHs have been isolated and characterized. However, the microbial degradation of PAHs in sediments is impaired due to the anaerobic conditions that result from the typically high oxygen demand of the organic material present in the soil, the low solubility of oxygen in water, and the slow mass transfer of oxygen from overlying water to the soil environment. For these reasons, anaerobic microbial degradation technologies could help alleviate sediment PAH contamination and offer significant advantages for cost-efficient in-situ treatment. But very little is known about the potential for anaerobic degradation of PAHs in field soils. The objectives of this research were to assess: (1) the potential for biodegradation of PAH in field aged soils under denitrification conditions, (2) to assess the potential for biodegradation of naphthalene in soil microcosms under denitrifying conditions, and (3) to assess for the existence of microorganisms in field sediments capable of degrading naphthalene via denitrification. Two kinds of soils were used in this research: Harbor Point sediment (HPS-2) and Milwaukee Harbor sediment (MHS). Results presented in this seminar indicate possible degradation of PAHs in soil under denitrifying conditions. During the two months of anaerobic degradation, total PAH removal was modest probably due to both the low availability of the PAHs and competition with other more easily degradable sources of carbon in the sediments. For both Harbor Point sediment (HPS-2) and Milwaukee Harbor sediment (MHS), PAH reduction was confined to 3- and 4-ring PAHs. Comparing PAH reductions during two months of aerobic and anaerobic biotreatment of MHS, it was found that extent of PAHreduction for anaerobic treatment was compatible with that for aerobic treatment. Interestingly, removal of PAHs from sediment particle classes (by size and density) followed similar trends for aerobic and anaerobic treatment of MHS. The majority of the PAHs removed during biotreatment came from the clay/silt fraction. In an earlier study it was shown that PAHs associated with the clay/silt fraction in MHS were more available than PAHs associated with coal-derived fraction. Therefore, although total PAH reductions were small, the removal of PAHs from the more easily available sediment fraction (clay/silt) may result in a significant environmental benefit owing to a reduction in total PAH bioavailability. By using naphthalene as a model PAH compound, biodegradation of naphthalene under denitrifying condition was assessed in microcosms containing MHS. Naphthalene spiked into MHS was degraded below detection limit within 20 days with the accompanying reduction of nitrate. With repeated addition of naphthalene and nitrate, naphthalene degradation under nitrate reducing conditions was stable over one month. Nitrite, one of the intermediates of denitrification was detected during the incubation. Also the denitrification activity of the enrichment culture from MHS slurries was verified by monitoring the production of nitrogen gas in solid fluorescence denitrification medium. Microorganisms capable of degrading naphthalene via denitrification were isolated from this enrichment culture.

• Journal of Life Science
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• v.21 no.11
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• pp.1599-1606
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• 2011

To evaluate the effects of microorganism agents on oil biodegradation, treatability and microcosm studies were conducted. Petroleum oil degrading bacteria were isolated from enriched cultures of oil-contaminated sediment samples using a mineral salts medium (MSM) containing 0.5% Arabian heavy crude oil as the sole carbon source. After a 5 day-incubation period using MSM, mixed microorganisms of three species (strains BS1, BS2 and BS4) degraded 48.4% of aliphatic hydrocarbons and 30.5% of aromatic hydrocarbons. Treatability and microcosm tests were performed in the three different treatment conditions (AO: Arabian heavy crude oil, AO+IN: Arabian heavy crude oil+inorganic nutrient, AO+IN+MM: Arabian heavy crude oil+inorganic nutrient+mixed microorganism agents). Among these, significantly enhanced biodegradation of aliphatic hydrocarbons were observed in AO+IN and AO+IN+MM conditions, without showing any different biodegradation rates in either condition. However, the degradation rates of aromatic hydrocarbons in an AO+IN+MM condition were increased by 50% in the treatability test and by 13% in the microcosm test compared to those in an AO+IN condition. Taken together, it can be concluded that mixed microorganism agents enhance the biodegradation of aliphatic and aromatic hydrocarbons in laboratory, a treatability test, and a microcosm test. This agent could especially be a useful tool in the application of bioremediation for removal of aromatic hydrocarbons.

• Proceedings of the Korean Institute of Navigation and Port Research Conference
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• 2016.05a
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• pp.132-134
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• 2016

The Study mainly focused on bioremediation of 16 types PAHs and heavy metals in contaminated marine sediments at filed scale study using slow release biostimulant ball (BSB) was investigated. In our experiment, filed scale study ($1m{\times}1m$) was performed and the effect of BSB on PAHs and heavy metals were analysed. BSB size and distance were determined and optimum size and distance were 3cm and 5.5cm respectively. BSB containing nutrients of acetate, nitrate and sulphate which can enhance the activity of microorganism to increase degrading capacity of PAHs and enhance the heavy metals stabilization also to decrease bioavailability. PAHs containing 16 types of 2, 3, 4, 5 and 6 rings compound PAHs were found and to degrade upto 100% in 2, 3 rings, upto 90 to 94% in 4 and 5 rings and 6 ring compound was degrade up to 70%. For heavy metals stabilization percentage was increased using BSB sediment and exchangeable portion was decreased and residual portion was increased in all analysed heavy metals. BSB enhance the PAHs degradation and stabilization of heavy metals percentages. BSB is a promising method for remediation of PAHs and heavy metals in contaminated marine sediments.

• Journal of Korea Soil Environment Society
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• v.3 no.2
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• pp.115-125
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• 1998

The biodegradation of ($C^{14}$)phenanthrene was studied in water and soil-water systems with nonionic surfactants and biosurfactant : polyoxyethylene alkyl ester($C_{17}$$H_33$COO($C_2$$H_4$O)nH) and sophorolipid. The extents of so1ubilization and biodegradation were monitored by radiotracer technique. Experimental results showed that surfactant concentrations above the critical micelle concentration were toxic to the phenanthrene-degrading bacteria in soil or active sludge and the presence of surfactant micelles inhibited mineralization of PAHs. Solubility and bioavailibility of phenanthrene in water and soil-water system were enhanced by mixed surfactants system. The optimum water content and hydrogen concentration were 30% (w/v) , pH 7, respectively.