오염된 토양으로부터 분리된 세균에 이한 2,4-Dinitrotoluene과 2,6-Dinitrotoluene의 생물학적 제거

오계현,조윤석,이명석 순천향대학교 기초과학연구소 1999 순천향자연과학연구 논문집 Vol.5 No.2

Dinitrotoluenes (e.g., 2.,4-Dinitrotoluene and 2,6-Dinitrotoluene) are the major impurity resulting from the manufacture of 2,4,6-trinitrotoluene(TNT) and are primary starting materials for the synthesis of toluenediisocyanate which are used in the production of polyurethane foam. Both 2,4-DNT(2,4-dinitro- toluene) and 2,6-DNT(2,6-dinitrotoluene) are recalcitrant and have been shown to be toxic to aquatic and terrestrial organisms in the environment. Because little information is available for the biological degradation of DNTs, they have has not been evaluated for the possible treatment processes of industrial waste streams where DNTs may represent a serious disposal problem. In the present work, we explored the feasibility using a bacterium for DNTs degradation, with the ultimate aim of application for the industrial treatment. A DNTs-degrading bacterium was isolated from soil samples collected from an explosive contaminated area. Complete degradation of 2,4-DNT at the initial concentration of 50 ㎎ 2,4-DNT per liter was achieved within 10 days of incubation in bench-scale bioreactors, whereas 2,6-DNT degraded completely within 14 days under the same conditions, respectively. The bacterium was able to utilize DNTs as the sole nitrogen source. Co-substrate was essential for the bacterial growth and DNTs degradation in the medium. Neither bacterial growth nor DNTs degradation were observed in the absence of co-substrate. The detection of the parent substrates and their intermediates were based on HPLC and GC-MS methodology.

오염된 토양으로부터 분리된 Burkholderia cepacia에 의한 2,4-D의 미생물학적 분해

오계헌,조윤석 순천향대학교 기초과학연구소 1999 순천향자연과학연구 논문집 Vol.5 No.1

Mixed cultures capable of utilizing 2,4-D (2,4-dichlorophenoxyacetic acid) as the sole carbon and energy source were enriched from contaminated soils which had a previous history of annual 2,4-D application. Among six strains isolated from the mixed cultures, the isolate which had an excellent 2,4-D degradability was selected. Morphological observation and various physiological characterization were performed for the isolate. MIS (microbial identification system) was used for the identification of the isolate, and assigned as Burkholderia cepacia. Complete degradation of 2,4-D was achieved in the experiment within 28 hours of incubation. 2,4-DCP(2,4-dichlorophenol) was produced as a metabolite during 2,4-D degradation. For the analyses of 2,4-D and 2,4-DCP, HPLC and UV-spectrophotometer were used in this study.

플라스크와 발효조에서 세균혼합배양에 의한 페녹시계 제초제 MCPP의 분해

오계헌 순천향대학교 1992 논문집 Vol.15 No.1

제초제 생산공장 주변으로부터 MCPP[2-(2-methyl-4-chlorophenoxy) propionic acid]를 이용하는 세균배양을 얻었다. UV-흡광법을 이용하여 플라스크와 발효조에서, 배양액의 잔존 MCPP 농도를 분석하였다. 잔존 MCPP 농도와 무기염소의 형성에 근거한 mass blance가 만들어졌다. 배양액에 유리되는 무기염소의양은 MCPP농도의 변화로부터의 기대치와 일치하였다. MCPP는 본 실험조건하에서 완전 분해되지 않았다. 세포단백질은 실험기간동안 배양액에 유리되는 무기염소의 농도에 비례하였다. MCPP분해효율과 분해율은 발효조에서 약 96%와 560 μmol/l·day 그리고 플라스크에서 약 50%와 140 μmol/l·day였다. Soil samples were collected from a fertilizer manufacturing plant site and used for enrichment of bacterial cultures with 2-(2-methyl-4-chlorophenoxy) propionic acid(MCPP) as the sole carbon and energy source. UV-spectrometry was used to analyze the residual MCPP concentration in the test cultures growing in shake flasks and a stirred tank reactor. A mass balance was established based upon the residual concentration of MCPP and formation of inorganic chloride in the test cultures. The amount of inorganic chloride released into solution was in agreement with the expected value calculated from the changes in MCPP concentration. MCPP was not completely degraded under the experimental condition. Cell protein paralleled a similar time profile to that of the inorganic chloride during the experiment. The efficiency and rate of MCPP degradation were approximately 96% and 560 μmol/l·day in the stirred tank reactor, and approximately 50% and 140 μmol/l·day in shake flasks.

컬럼반응조내에서 과립 활성탄상에 고정된 미생물에 의한 BTX(벤젠-톨루엔-크실렌)의 제거에 관한 연구

오계헌,윤경하 순천향대학교 부설 산업기술연구소 1995 순천향 산업기술연구소논문집 Vol.1 No.2

The microbiological degradation of benzene(B), toluene(T), and p-xylene(X) was studied in column reactors with granular activated carbon(GAC) as a support matrix. For the initial experiment, bildegradation of B, T and X was evaluated in the test cultures growing in shake-flasks, respectively. Microbial growth was measured by dry weight and degradation of BTX was verified by HPLC, GC and UV/vis spectroscopic analysis of the residual substrates concentration in the test cultures. The effects on the degradation of B, T and X by pH, substrate concentration and supplemented ascorbic acid were compared. Immobilization of BTX-degrading bacteria on GAC was examined by scanning electron microscope. The test culture was stabilized as a fixed-film system in the GAC column reactor. BTX degradation was evaluated with a batch and continuous mode of operation of the GAC fixed-film column reactors. Concurrent degradation occurred on mixtures of benzene, toluene and p-xylene.

TNT-분해세균인 Stenotrophomonas maltophilia에 의한 s-Triazine계 제초제인 Atrazine의 미생물학적 분해

오계헌,조윤석,이명석 순천향대학교 기초과학연구소 1999 순천향자연과학연구 논문집 Vol.5 No.2

The agricultural use and relative persistence of s-triazine herbicide such as atrazine have led to increasing concern about environmental contamination. Stenotrophomonas maltophilia capable of utilizing TNT as sole nitrogen source has been isolated from contaminated soils and the bacterium showed excellent degradability for several-triazine herbicides including atrazine. Complete depletion of atrazine was achieved within 30 hours of incubation. Atrazine was catabolized in the presence of different supplemented carbons. Among the co-substrates used in this experiment (e.g., glucose, fructose, starch, succinate, acetate), fructose was the best co-substrate. However, no atrazine degradation was monitored without the supplemented co-substrates in the cultures. The relationships between atrazine degradation by S. maltophilia and several relevant physicochemical parameters (e.g., N-sources, yeast extract) were examined. High performance liquid chromatographic methodology was used to measure this substrate and it also resolved unknown intermediates.

Pseudomonas putuda Hk-6에서 2,4,5-Trinitroroluene에 의한 스트레스 충격 단백질의 생성

장효원,오계헌 순천향대학교 기초과학연구소 2000 순천향자연과학연구 논문집 Vol.6 No.2

The aim of this work was to examine the production of stress shock proteins by 2,4,6-trinitrotoluene (TNT) in Pseudomonas putida HK-6 which isolated from explosive contaminated sites. The stress shock proteins, which contribute to the resistance of the cytotoxic effect of TNT, were induced at various concentrations of TNT in exponentially growing cultures of P. putida HK-6. Theses responses involved the induction of a 70-kDa DnaK and a 60-kDa GroEL proteins, characterized by SDS-PAGE and Western blot by use of anti-DnaK and anti-GroEL monoclonal antibodies. Survival of P. putida HK-6 with time in the presence of different concentrations of TNT was monitored, and viable counts paralleled the induction of the stress shock proteins in this strain.

TNT 분해세균 Pseudomonas sp. HK-6에서 정제한 NAD(P)H-nitroreductase의 특성연구

호은미,오계현 순천향대학교 기초과학연구소 2003 순천향자연과학연구 논문집 Vol.9 No.2

The aim of this work was to characterize NAD(P)H-nitroreductase purified from Pseudomonas sp. HK-6, which could utilize 2,4,6-trinitrotoluene. In initial experiments, NAD(P)H-nitroreductase from actively growing HK-6 cells was separated by ammonium sulfate precipitation, DEAE-sepharose, and Q-sepharose. Specific activity of NAD(P)H-nitroreductase was approximately 4.46 unit/㎎. Optimum temperature and pH of the enzyme were approximately 30℃ and 7.5, respectively. Specific activity of NAD(P)H-nitroreductase on the effect of several metal ions was monitored. Fe^(3+) showed stimulatory effect on the activity of NAD(P)H-nitroreductase, whereas the specific activity of the enzyme was determined to approximately 27 kDa by SDS-PAGE.

토양에서 분리된 Acrylamide 분해 세균 JK-7의 분리 및 특성

천재우,오계현 순천향대학교 기초과학연구소 2003 순천향자연과학연구 논문집 Vol.9 No.2

The feasibility of using pure culture for acrylamide degradation, with the ultimate aim of application for biological treatment process, was explored. The present study reports on the test cultures which were developed to grow aerobically with acrylamide as the sole source carbon and nitrogen. A bacterial isolate, strain JK-7 was isolated from paddy soil samples, Strain JK-7 could degrade 50 mM acrylamide completely within 72 hours of incubation. Major intermediates resulting from acrylamide degradation were not detected with the HPLC methodology except acrylic acid which appeared to accumulate transiently in the growth medium. Initial pH 7.2 of the cultures became to increase 8.4 at the end of incubation. When JK-7 cells were grown at over 100 mM acrylamide, there was a pause of cell growth, resulting in a reduction in the rates of acrylamide degradation. Survival test revealed that cells exposed to low concentrations of acrylamide enable to the strain JK-7 to survive at a lethal concentration of 100 mM acrylamide. The relationships between the acrylamide degradation by JK-7 and several relevant physicochemical environmental parameters were examained. The effect of supplemented carbons(e.g., glucose, fructose, citrate, succinate) on the acrylamide degradation by the test culture of JK-7 was evaluared. The results indicated that the addition of carbons accelerated the bacterial growth and acrylamide degradation compared to in the absence of supplemented carbons. The effect of supplemented nitrogens on the degradation was monitored. Increasing concentrations of yeast extract resulted in higher growth yield, based on the turbidity measurement, and complete degradation of acrylamide. However, acrylamide degradation was essentially uninfluenced by the addition of (NH_(4))_(2)So_(4),NH_(4)Cl, or urea. The bacterium was identified as belonging to the genus Pseudomonas in the basis of use BIOLOG test, and designated as Pseudomonas sp. JK-7

Delftia sp. JK-2로부터 분리된 Catechol 2,3-dioxygenase의 정제와 특성 연구

황선영,오계헌 순천향대학교 기초과학연구소 2002 순천향자연과학연구 논문집 Vol.8 No.2

The purpose of this work was to investigate the purification and characterization of catechol 2,3-dioxygenase isolated from Delftia sp. JK-2, which could utilize aniline as the sole source of carbon, nitrogen and energy. In initial experiments, several characteristics of C2,3O separated with ammonium sulfate precipitation and DEAE-sepharose were investigated. Specific activity of C2,3O was approximately 4.72 unit/㎎. C2,3O demonstrated its enzyme activity to another substrates including catechol and 4-methylcatechol. The optimum temperature of C2,3O was 30℃, and the optimal pH was approximately 8. Metal ions including Ag^(+), Hg^(+), and Cu^(+2) showed inhibitory effect on the activity of C2,3O. Molecular weight of the enzyme was determined to approximately 35 kDa by SDS-PAGE.

Benzoate에서 배양된 Acinetobacter baumannii로부터 Catechol 1,2-dixoygenase의 정제와 촉매적 성질

송승열,오계헌 순천향대학교 기초과학연구소 2001 순천향자연과학연구 논문집 Vol.7 No.2

The purpose of this work was to perform the catalytic properties and sequence analysis of catechol 1,2-dioxygenase (C1,2O) purified from Acinetobacter baumannii which was grown on benzoate as a carbon source. C1,20 demonstrated its enzyme activity to other substrates, catechol and 4-methylcatechol. The optimum temperature of C1,20 was 35℃, and the optimal pH was in the range from pH 7.5 to 9. Ag^+, Hg^+, and Cu^+2 showed inhibitory effect on the activity of C1,20. Molecular weight of the enzyme was determined to approximately 36 kDa by SDS-PAGE. N-terminal amino acid sequence of C1,20 was analysed as ^1MNYQQIDALVKQMNVDTAKG^20 and exhibited 95% sequence homology with that of C1,20 from Acinetobacter radioresistens. For internal sequencing analysis, trypsin digestion and peptide mapping were performed. Molecular weights of three digested peptide fragments were analyzed as 966.3 dalton, 1933.8 dalton and 2081.7 dalton by MALDI-TOF, which were matched with internal sequences (^1SQSDFNLRR^9, ^1TIEGPLYVAGAPESVGFAR^19, ^1HGNRPSHVHYFVSAPGYR^18)of Acinetobacter radioresistens.