• 한국지하수토양환경학회지:지하수토양환경
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• 제21권1호
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• pp.72-79
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• 2016

Bioremediation is one of the most effective ways to remediate TPH-contaminated sites. However, under actual field conditions that are not at the optimum temperature, degradation of microorganisms is generally reduced, which is why the efficiency of biodegradation is known to be significantly affected by the soil temperature. Therefore, in this study, the labscale experiment was conducted using indigenous crude oil degrading microorganisms isolated from crude oil contaminated site to evaluate the remediation efficiency. Crude oil degrading microorganisms were isolated from crude oil contaminated soil and temperature, which is a significant factor affecting the remediation efficiency of land farming, was adjusted to evaluate the microbial crude oil degrading ability, degradation time, and remediation efficiency. In order to assess the field applicability, the remediation efficiency was evaluated using crude oil contaminated soil (average TPH concentration of 10,000 mg/kg or more) from the OO premises. Followed by the application of microorganisms at 30℃, the bioremediation process reduced its initial TPH concentration of 10,812 mg/kg down to 1,890 mg/kg in 56 days, which was about an 83% remediation efficiency. By analyzing the correlation among the total number of cells, the number of effective cells, and TPH concentration, it was found that the number of effective microorganisms drastically increased during the period from 10 to 20 days while there was a sharp decrease in TPH concentration. Therefore, we confirmed the applicability of land farming with isolated microorganisms consortium to crude oil contaminated site, which is also expected to be applicable to bioremediation of other recalcitrant materials.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2001년도 추계학술발표대회
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• pp.578-581
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• 2001

A treatability study was conducted using a hydrocarbon-contaminated soil for the oPtimization of bioremediation strategy best fit to a given set of contamination. The applicability of nutrients, biosurfactant, and oil-degrading microorganisms were examined by monitoring $CO_2$ evolution and oil degradation The addition of inorganic nutrients in the form of slow released fertilizer accelerated the initial rate of $CO_2$ evolution by a factor of 3. The application of oil-degrading microorganisms did not significantly increased $CO_2$ evolution or biodegradation efficiency. Application of a commercial biosurfactant was most effect in terms of the total $CO_2$ evolution and the oil degradation rate. The results indicate that $CO_2$ evolution measurement was found to be a simple and reliable countermeasure of crude oil hydrocarbon mineralization for the rapid determination of the best-fit bioremediation strategy.

• Journal of Microbiology
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• 제37권3호
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• pp.128-135
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• 1999

Among oil-degrading microorganisms isolated from oil-polluted industrial areas, one yeast strain showed high degradation activity of aliphatic hydrocarbons. From the analyses of 18S rRNA sequences, fatty acid, coenzyme Q system, G+C content of DNA, and biochemical characteristics, the strain was identified as Yarrowia lipolytica 180. Y. lipolytica 180 degraded 94% of aliphatic hydrocarbons in minimal salts medium containing 0.2% (v/v) of Arabian light crude oil within 3 days at 25$^{\circ}C$. Optimal growth conditions for temperature, pH, NaCl concentration, and crude oil concentration were 30$^{\circ}C$, pH 5-7, 1%, and 2% (v/v), respectively. Y. lipolytica 180 reduced surface tension when cultured on hydrocarbon substrates (1%, v/v), and the measured values of the surface tension were in the range of 51 to 57 dynes/cm. Both the cell free culture broth and cell debris of Y. lipolytica 180 were capable of emulsifying 2% (v/v) crude oil by itself. They were also capable of degrading crude oil (2%). The strain showed a cell surface hydrophobicity higher than 90%, which did not require hydrocarbon substrates for its induction. These results suggest that Y. lipolytica has high oil-degrading activity through its high emulsifying activity and cell hydrophobicity, and further indicate that the cell surface is responsible for the metabolism of aliphatic hydrocarbons.

• Journal of Microbiology and Biotechnology
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• 제10권3호
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• pp.333-337
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• 2000

An oil-degrading yeast, Yarrowia lipolytica 180, exhibits interesting cell surface characteristics under the growth on hydrocarbons. An electron microscopic study revealed that the cells grown on crude oil showed protrusions on the cell surface, and thicker periplasmic space and cell wall than the cell surface, and thicker periplasmic space and cell wall than the cells grown on glucose. Y. lipolytica cells lost its cell hydrophobicity after pronase(0.1 mg/ml) treatment. The strain produced two types of emulsifying materials during the growth on hydrocarbons; one was water-soluble extracellular materials and the other was cell wall-associated materials. Both emulsifying materials at lower concentration (0.12%) enhanced the oil-degrading activity of Moraxella sp. K12-7, which had medium emulsifying activity and negative cell hydrophobicity; however, it inhibited the oil-degrading activity of Pseudomunas sp. K12-5, which had medium emulsifying activity and cell hydrophobicity. These results suggest that the oil-degrading activity of Y. lipolytica 180 is closely associated with cell surface structure, and that a finely controlled application of Y.lipolytica 180 in combination with other oil-degrading microorganisms showed a possible enhancing efficiency of oil degradation.

• 생명과학회지
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• 제24권6호
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• pp.694-699
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• 2014

토착 미생물은 친환경적 정화에 중요한 역할을 한다. 원유(crude oil)를 분해하는 80균주를 인천 연안에서 분리하고 oil film collapsing방법을 이용하여 유화능이 있는 12균주를 선별하였다. 이들 균주에 대해 ${\rho}$-nitrophenyl butylrate를 기질로 이용하여 리파아제(lipase)활성과 n-hexanedecane을 기질로 이용하여 유화(emulsification)활성을 측정하여 원유 분해 활성이 좋은 Incheon9를 선별하고 gas chromatography (FID)로 paraffine계 탄화수소를 감소시키는 것을 확인하였다. 이 균주의 16s rRNA유전자 분석을 통해 Acinetobacter sp.로 동정하고 NCBI에 등록하여 accession code (KF548540)를 부여 받았다. Acinetobacter sp. Incheon9의 성장과 유화능이 최적 배양 조건은 $20^{\circ}C$, pH 7, 1% NaCl였으며 대수증식기 기간에 가장 높은 유화능을 보였으며 탄화수소가 짧은 trybutyrin에서 분해능력이 좋았다. 이번 연구결과는 환경오염에 활용 가능한 미생물자원군의 확보를 위한 연구였으며 추후 활용을 위해서는 실제 환경에서 동일한 활성을 가지는지 여부에 대한 연구가 추가로 진행되어야 할 것이다.

• 미생물학회지
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• 제34권3호
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• pp.101-107
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• 1998

지속성영양염제(Slow Release Fertilizer, SRF)와 유류분해미생물을 이용한 생물학적 방법으로 오염된 자갈환경으로부터 유류를 제거하는 방법을 연구하기 위해 microcosm 실험을 수행하였다. Microcosm은 직경 40 mm 이하의 자갈을 직경 10 cm 높이 20 cm의 glass column에 충진하여 준비하였으며, 전 실험구에 유류 약 2.4%(w/v)를 첨가하였다. 실험구 I은 대조구로서 아무것도 첨가되지 않았으며, 실험구 II에는 Inipol EAP-22(Elf Aquitaine, France)와 미생물제제, 실험구 III에는 SRF와 미생물제제, 실험구 IV에는 Inipol EAP-22가 첨가되었다. 유류의 생물학적 분해도를 $C_{17}$/Pristane과 $C_{18}$/Phytane 비율료 나타내었을 때, 실험구 III에서 유류분해도가 가장 높을 것으로 나타났다. 종속영양미생물 및 유유분해미생물의 전체적인 변화와 전자전달계 활성도는 유류분해도의 변화와 유사한 양상을 보이며 변화하는 것으로 나타났다. 무기영양염류의 분석 결과 실험구 III 에서는 전 실험기간에 걸쳐 질소와 인이 제한요인으로 작용하지 않은 것으로 나타났으며, 이는 SRF가 지용성영양염제인 Inipol EAP-22에 비해 지속적으로 영양염류를 제공함으로써 자갈환경에서 유류분해가 증가되었다고 추정할 수 있다. 따라서 본 실험결과로부터 SRF와 유류분해 미생물제제를 사용하여 해안 자갈지역의 유류오염을 효율적으로 제거할 수 있는 가능성이 확인되었다.

• 생명과학회지
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• 제21권11호
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• pp.1599-1606
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• 2011

유류분해에 있어 혼합미생물제제의 효과를 평가하기 위해 미생물제제의 처리성능과 microcosm test를 수행하였다. 유류분해세균은 0.5% Arabian heavy crude oil을 유일 탄소원으로 제공된 최소배지를 이용한 연속적인 농후배양을 통하여 분리하였다. 우수 유류분해 미생물조합인 3종의 균주(BS1, BS2, BS4)는 MSM배지에서 5일의 배양기간 동안 지방족 탄화수소를 48.4%, 방향족 탄화수소를 30.5% 생분해하였다. 처리성능 및 microcosm test는 Arabian heavy crude oil을 첨가한 후 3가지 처리조건인 무처리, 무기영양염처리 그리고 무기영양염 및 혼합미생물처리조건에서 유류화합물의 생물분해에 미치는 영향을 조사하였다. 무기영양염처리구와 무기영양염 및 혼합미생물처리구에서 지방족 탄화수소의 분해율은 실험기간 동안 유의하게 향상되었으며 두 실험구간 유의한 차이는 관찰되지 않았다. 그러나 무기영양염 및 혼합미생물처리구에서 방향족 탄화수소의 생분해율은 무기영양염제만을 처리한 시험구와 비교하여 처리성능 시험의 경우 50% 그리고 microcosm test의 경우 13%를 향상시켰다. 본 연구의 결과로부터 혼합미생물제제는 실험실, 처리성능 및 microcosm test에서 지방족뿐만 아니라 방향족 탄화수소의 생물분해를 촉진하였다. 특히 혼합미생물제제는 방향족 탄화수소의 제거를 위한 생물정화기술의 적용에 있어 유용한 도구로 판단된다.

• Journal of Microbiology
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• 제44권3호
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• pp.354-359
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• 2006

This study examined the capacity of immobilized bacteria to degrade petroleum hydrocarbons. A mixture of hydrocarbon-degrading bacterial strains was immobilized in alginate and incubated in crude oil-contaminated artificial seawater (ASW). Analysis of hydrocarbon residues following a 30-day incubation period demonstrated that the biodegradation capacity of the microorganisms was not compromised by the immobilization. Removal of n-alkanes was similar in immobilized cells and control cells. To test reusability, the immobilized bacteria were incubated for sequential increments of 30 days. No decline in biodegradation capacity of the immobilized consortium of bacterial cells was noted over its repeated use. We conclude that immobilized hydrocarbon-degrading bacteria represent a promising application in the bioremediation of hydrocarbon-contaminated areas.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.435-438
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• 2000

토양으로부터 2종류 그리고 해양으로부터 1종류의 원유 분해 미생물을 분리하였고, 이들을 strain A132, strain F722 그리고 strain OM1으로 각각 명명하였다. 이들 미생물은 Acinetobacter sp., Pseudomonas aeruginosa, Acinetobacter calcoaceticus로 각각 동정되었다. Strain A132. F722의 최적 배양 및 분해온도는 $35^{\circ}C$ 이고, 최적 생장 pH는 각각 8과 9에서 나타났다. 원유를 유일한 탄소원으로 하여 배양을 하였을 때 원유농도 2.0%(w/v)에서 생장이 높았다. 한편, 해양에서 분리된 strain OM1은 pH 7, 원유농도 3.0%(w/v)에서 생장이 높았다. 원유 분해능 조사에서는 Eleuthera (OMAN) 원유를 2.0%(w/v) 기질로 하였을 때, strain A132가 $25^{\circ}C$에서 $5.49g/\;l\;{\cdot}\;day$의 분해능을 나타내었다. 반면, L-Zakum (AFRICA) 원유에서는 strain F722가 $35^{\circ}C$에서 $1.19g/\;l\;{\cdot}\;day$의 분해능을 보였다. 등유$(nC_9-nC_{20})$와 경유$(nC_9-nC_{28})$에 대하여 분해특성을 조사한 결과 strain OM1은 배양 7일 후 95, 75%를 각각 분해하였다. Strain F722는 배양10일 후 80%의 분해율을 나타내었다.

• Journal of Microbiology and Biotechnology
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• 제22권9호
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• pp.1185-1192
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• 2012

This study investigated the hydrocarbonoclastic microbial community present on weathered crude oil and their ability to degrade weathered oil in seawater obtained from the Gulf St. Vincent (SA, Australia). Examination of the native seawater communities capable of utilizing hydrocarbon as the sole carbon source identified a maximum recovery of just $6.6{\times}10^1\;CFU/ml$, with these values dramatically increased in the weathered oil, reaching $4.1{\times}10^4\;CFU/ml$. The weathered oil (dominated by > $C_{30}$ fractions; $750,000{\pm}150,000mg/l$) was subject to an 8 week laboratory-based degradation microcosm study. By day 56, the natural inoculums degraded the soluble hydrocarbons (initial concentrations $3,400{\pm}700mg/l$ and $1,700{\pm}340mg/l$ for the control and seawater, respectively) to below detectable levels, and biodegradation of the residual oil reached 62% ($254,000{\pm}40,000mg/l$) and 66% ($285,000{\pm}45,000mg/l$) in the control and seawater sources, respectively. In addition, the residual oil gas chromatogram profiles changed with the presence of short and intermediate hydrocarbon chains. 16S rDNA DGGE sequence analysis revealed species affiliated with the genera Roseobacter, Alteromonas, Yeosuana aromativorans, and Pseudomonas, renowned oil-degrading organisms previously thought to be associated with the environment where the oil contaminated rather than also being present in the contaminating oil. This study highlights the importance of microbiological techniques for isolation and characterisation, coupled with molecular techniques for identification, in understanding the role and function of native oil communities.