Aspergillus 속에서의 Amylase 및 Protease의 고생산 역가를 가진 균주의 개발

玄馨煥 한국외국어대학교 1995 한국외국어대학교 논문집 Vol.28 No.2

Trichoderma viride 의 변이균주 분리를 위한 Cellulase Semiquantitative Plate Assay

현형환,백형석,이인복,이세영 ( Hyung Hwan Hyun,Hyung Suk Baik,In Bok Lee,Se Yong Lee ) 한국미생물생명공학회 1978 한국미생물·생명공학회지 Vol.6 No.3

Production of Acrylamide Using Immobilized Cells of Rhodococcus rhodochrous M33

현형환,Bu-Youn Kim 한국생물공학회 2002 Biotechnology and Bioprocess Engineering Vol.7 No.4

The cells of Rhodococcus rhodochrous M33, which produce a nitrile hydratase enzyme, were immobilized in acrylamide-based polymer gels. The optimum pH and temperature for the activity of nitrile hydratase in both the free and immobilized cells were 7.4 and 45oC, respectively, yet the optimum temperature for acrylamide production by the immobilized cells was 20oC. The nitrile hydratase of the immobilized cells was more stable with acrylamide than that of the free cells. Under optimal conditions, the final acrylamide concentration reached about 400 g/L with a conversion yield of almost 100% after 8 h of reaction when using 150 g/L of immobilized cells corresponding to a 1.91 g-dry cell weight/L. The enzyme activity of the immobilized cells rapidly decreased with repeated use. However, the quality of the acrylamide produced by the immobilized cells was much better than that produced by the free cells in terms of color, salt content, turbidity, and foam formation. The quality of the aqueous acrylamide solution obtained was found to be of commercial use without further purification.

호열 혐기성 세균의 산업적 이용 가능성

현형환 한국미생물학회 1988 微生物과 産業 Vol.14 No.2

산학협동위원회 보고 : 우리의 치즈는 ?

현형환 한국미생물생명공학회 ( 구 한국산업미생물학회 ) 2001 생물산업 Vol.14 No.4

Fermentation Conditions for the Production of Cell Mass in Streptococcus faecalis and Endospores in Bacillus mesentericus

현형환,김부연,류한봉 한국외국어대학교 외국학종합연구센터 부설 기초과학연구소 1996 기초과학연구 Vol.4 No.-

Streptococcus faecalis는 glucose, lactose등을 탄소원으로 잘 이용하였으며 glucose를 탄소원으로, yeast extract와 casitone을 질소원으로 구성된 배지를 사용하여 pH 7.0에서 control되는 fermentor 배양에서 optical density(660nm) 7.3(viable cell concentration 6.5×10(9) cells/㎖)이상의 높은 cell concentration을 얻을 수 있었다. Bacillus mesentericus의 sporulation을 최대화 하기 위한 발효 조건을 조사한 결과 glucose를 위한 최적 CaCl₂농도는 0.lmM이었다. Streptococcus faecalis rapidly fermented glucose and lactose as a C-source, and the maximum cell concentration was obtained when grown in a jar fermentor that contained YCG medium consisting of glucose as a C-source and casitone and yeast extracts as a N-source, and that was controlled at pH 7.0. From experiments to maximize the sporulation efficiency in Bacillus mesentericus, glucose caused the decrease in sporulation efficiency, and calcium chloride was stimulatory for sporulation, of which optimum concentration was 0.lmM.

Salmonella/microsomal enzyme activation system에서의 농약의 돌연변이 유발성

변우현,현형환,이세영 한국미생물학회 1976 미생물학회지 Vol.14 No.3

19 pesticides including 12 insecticides, 2 herbicides and 5 fungicides have been tested for mutagenic activity in the Salmonella/microsome system. It was found that insecticides, DDVP, Trichlorfon, Sumithion, Naled, fungicide, TMTD and herbicide NIP induced base substitute mutation and herbicide MO frameshift mutation. Mutagenicity of Sumithion and NIP was appeared only after rat microsomal enzyme activation and that of TMTD was increased after the microsomal enzyme activation.

Streptomyces. orientalis에 의해 생산된 vancomycin의 정제

정형무,현형환 한국외국어대학교 기초과학연구소 2001 기초과학연구 Vol.11 No.-

vancomycin은 glycopeptide계 항생제로서 세포벽에 작용하기 때문에 외부로 분비되지만 일부는 자신의 세포벽에 결합되어 있다 따라서 발효액으로부터 균체를 제거할 때 vancomycin 회수율을 향상시키기 위해 발효액의 pH 및 회석배율에 따른 vancomycin 회수율을 조사한 결과, 발효액을 10배 이상 희석하고 pH 2.0으로 보정하여 균체를 제거 하였을 때 가장 높은 vancomycin 회수율을 나타내었다. 그러나 전체적인 정제 공정을 고려하여 2-3배 희석하고 pH 2.0으로 보정할 때 발효원액 자체를 여과할 때보다 75% 높은 회수율을 나타냈다. Vancomycin을 정제하기 위해 강산성 양이온 교환수지인 Deuolite C20과 약염기성 음이온 교환 수지인 Deuolite A56l 및 강염기성 음이온 교환수지 Diaion PA406에 대하여 탈색, 순도 및 수율을 조사하였다. 탈색, 수율 및 순도에서 C20(Na+)과 PA406(Cl-)이 효과적이었다. 양이온 교환 수지 및 음이온 교환 수지를 통과한 액을 비이온성 흡착수지 HP-20에 통액하여 vancomycin을 흡착시킨 후, 30% 메탄올로 용출하였을 때, 순도 94%의 vancomycin을 수율 77.5%로 분리할 수 있었다. HP20으로부터 분리한 vancomycin 분획에 남아 있는 color물질 및 pyrogen을 제거하기 위해 vancomycin 활성분획을 alumina에 통액한 결과, 발효 여액대비 수율은 90%이었고 순도는 91%로 증가하였으며 endotoxin은 0.312EU/mg-vancomycin 이었다. Alumina의 vancomycin 분획을 50mg/ml로 농축하여 CM-Toyopearl(NH4+)에 통액하고 0.5N HCl로 vancomycin을 용출시킨 결과, 발효 여액에 대한 수율은 87%였고 순도 99%까지 향상시킬 수 있었으며 endotoxin은 0.184EU/mg-vancomycin로 감소하였다. Vancomycin 결정을 얻기 위해 농축 농도 및 온도에 대하여 조사한 결과, 80mg/ml 이상 농축하여 30-40℃에서 결정화하였을 때 수율 및 순도가 가장 높았다. 따라서 CM-Toyopearl의 vancomycin 활성분획을 80mg/ml로 농축하고 10% NaOH로 pH8.2가 되도록 조정한 후, 30℃에서 1시간 교반하여 vancomycin을 석출시켰다. 석출된 vancomycin은 여과, 세정하여 건조한 결과 최종 정제 수율은 64%였으며 순도는 99.4%이었으며 endotoxin은 0.180EU/mg 이었다. Purification of vancomycin Ion exchange resins, DC-20, DA-561 and PA-406 were applied for discoloration and improvement of purity and yield in vancomycin purification. Cation exchange resin DC-20(Na+) and anion exchange resin DA-561 (Cl-) appeared to be most efficient for pre-treatment of culture supernatant for purification of vancomycin. The filtrate passed through the ion exchange columns, DC-20(Na+) and DA-56(Cl-) was then loaded on a nonfunctional type adsorption column HP20 and eluted with 30% methanol. of which the yield and purity was 94% and 77.5% respectively. In order to remove the color impurities of HP20 active fractions, active carbon and alumina were applied, and as the result, alumina was more efficient in terms of decoloration and purification yield than active carbon. When the active fractions passed through the HP20 column were treated with alumina. The purity was improved from 77.5% to 91% with 90% of yield, CM-toyopearl 650M (NH4+), a cation exchange resin was used as a next adsorption column to increase the purity of vancomycin. When the active fractions passed through alumina column were loaded on a CM-toyopearl column at the flow rate of 1.5bv/h, after concentrating to 50mg/ml of vancomycin using a reverse osmosis membrane system, the yield and purity was 87% and 99%, respectively. Optimum concentration and temperature for crystallization of vancomycin were also investigated. Then, the active fractions of CM-toyopearl column were collected and concentrated to 80mg/ml of vancomycin for crystallization. The highest yield and purity were obtained when crystallization was conducted at pH 8.2 and 30-40℃. By a series of purification processes, the crystallized vancomycin with 99.4% purity was obtained at 64% yield.

Cephalosporium acremonium var. M-10에 의한 Cephalosporin C 생산에 관한 연구

이호근,현형환,오두환,유주현 연세대학교 산업기술연구소 1990 논문집 Vol.22 No.2

The microorganism which produces Cepholsoporin C, has been selected from Cephalosporium acromonium by U.V radiation. Among the mutants, Cephalosporium acremonium var. M-10 was showed the highest Cephalosporin C productivity. On the basis of physiologieal characteristics, the difference hetween Cephalosporium acremonium var. M-10 and Cephalosporium acromonium was that the mutant did not assimilate sodium acetate, potassium nitrate, sodium nitrate. By using Cephalosporium acremonium var. M-10, the optimum conditions for the production of cephalosporin C on the rotary shaker were pH 7.0 soluble starch 4.0 w/v%, (NH₄)₂SO₄0.75w/v%,, CaCl ₂w/v%, soy bean meal 2w/v%, oleic acid 0.09w/v%, methionine 0.3w/v%, at 30℃ for 120 hours, respectively. On the study of time course, it revealed that the peak appeared at the stationery phese. The amount of Cephalosgorin C produced in this medium by mutant was about two times larger than that by Cephalosporium acremonium.

Optimization of fermentation conditions for the enhanced production of lacticin BH5

김미희,현형환 한국외국어대학교 외국학종합연구센터 부설 기초과학연구소 2003 기초과학연구 Vol.15 No.-

넓은 항균의 범위와 높은 활성을 나타내는 lacticin BH5의 생산 증대를 위한 최적 배양 조건을 조사하였다. shake-flask 배양시 lacticin BH5 생산을 위해서는 초기 배지 pH를 6.0으로 조절하여 배양하는 것이 적절하였으며, lacticin BH5의 생산에 있어서 최적 발효 배지를 조사한 결과, 탄소원으로는 glucose, 질소원으로 0.5% tryptone, 1% yeast extract 그리고 0.5% beef extract를 혼합한 배지에 1.25%의 K_2HPO_4와 무기염류로는 0.005% CuSO_4· 5H_2O를 복합적으로 첨가시킨 변형 MRS 배지에서 growth 및 박테리오신 생산이 최대였다. 또한 growth 및 박테리오신 생산에 대한 최적 온도는 30˚C인 것으로 나타났다.