CHO 세포 부유배양용 플라스크의 산소전달속도 모델 개발

고한철,유현욱,백종윤 한국생물공학회 2022 KSBB Journal Vol.37 No.1

Shake flasks are widely used as small-scale mammalian culture vessels during cell line development and bioprocess optimization for the production of biopharmaceuticals. However, the lack of information as well as controllability on dissolved oxygen often leads to oxygen limitation that can adversely affect cell growth and productivity during cell culture, thereby challenging cell line development and scale-up processes. In this work, we report an oxygen transfer rate (OTR) prediction model of shake flasks using Chinese hamster ovary (CHO) cells to avoid oxygen limitation issues in shake flasks. First, we varied shaking speed and culture volume to measure maximum OTR (OTR max ) and created an OTR max regression function. Then, we estimated theoretical maximum viable cell density with the specific oxygen uptake rate (sOUR) of CHO cells and the OTR max values. Lastly, we tested if decreasing culture volume or increasing shaker speed could mitigate the adverse effect of oxygen limitation. In conclusion, we believe this OTR prediction model would provide information about the oxygen availability in shake flasks, presumably assisting to achieve better culture performances.

Pseudomonas aeruginosa의 항생제내성에 관하여

高漢喆,朴政枰,朴悅 朝鮮大學校 附設 醫學硏究所 1989 The Medical Journal of Chosun University Vol.14 No.1

In order to study the MIC distribution and serotypes of p. aerugmosa, 112 strains isolated from two general hospital in kwangju city were tester). The following results were obtained. 1. Among 112 strains of p. aeruginosa isolated, 107 strains(95.5%) were typable. serotype E(30.4%) and G(30.4%) were most prevalent followed by B(11.6%), serotype A(7.1%) and serotype F, I(1.8%) among 107 typable strains. 2. According to test of MIC distribution on 17 antibiotics against p. aeruginosa. aminoglycosides were 0. 5~>512mcg/㎖, MIC peak in AK 32mcg/㎖ and other antimicrobial agents were > 512mcg/㎖. penicillins were 4~>512mcg/㎖, MIC peak in pi 256mcg/㎖ and other antimicrobial agents were >512mcg/㎖. Cephalosporines were 8~>512mcg/㎖, MIC peak in CP 64mcg/㎖ and other antimicrobial agents were>512mcg/㎖. 3. Among those antimicrobial agents, amikacin and rifampin showed good antimicrobial activity. 4. All of drug-resistant p. aeruginosa strains were multiply resistant to nine or more drugs. 35 strains were resistant to 17 drugs, follwed by 21 strains resistant to 15 drugs, 9 strains resistant to 14 drugs. 5. Out of 9~17 drugs, E. and G serotypes were 34 strains, out of 9~17 drugs, B sero-type were 13 strains, out of 13~]7 drugs, A serotype were 8 strains.

Pseudomonas aeruginosa의 항생제내성에 관하여

고한철 · 박정평 · 박열 조선대학교 의학연구원 1989 The Medical Journal of Chosun University Vol.14 No.1

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녹농균의 quinolone제재에 대한 감수성 및 혈청형 분포

최승호,고한철,이현국,박정평,임용,양남웅,김학렬,박열 朝鮮大學校 附設 醫學硏究所 1991 The Medical Journal of Chosun University Vol.15 No.1

123 strains of Pseudomonas aeruginosa isolated from various clinical specimens were tested for the distrution of serotypes and the evaluation of the antimicrobial susceptibility to each generation quinolone impounds. The results were as follows : 1. 107 strains(87%) were typable out of clinical isolate 123 strains. Serotype G was prevalent with 33 strains (26.8%) and followed by serotype E, 31 strains (25.2%), serotype B, 15 strains (12.2%) and successively typed in the order of serotype A, C, D, F, H, I, L, M, but serotype J,K and N were not entirely typed. 2. MIC peak was 1024ug/ml in case of nalidixic acid, 32ug/ml in pipemidic acid, 0.25ug/ml in ciprofloxacin, 2ug / ml in ofloxacin. 2ug/ml in enoxacin. 0.5ug/ml in norfloxacin, finally 8ug/ml in rosoxacin. 3. The second generation quinolone compound, pipemidic acid exhibited 8 to 30 times higher activity to Pseudomonas aeruyrtcisa than the first generation quinlolne, nalidixic acid. And the third generation compounds showed 4 to 100 times higher activities than the second generation quinolone. Especially ciprofloxacin showed 2 to 30 times higher antimicrobial activity than other generation quinolone compounds. 4. There were no significant differences to susceptibility patterns among the serotypes.

Thiobacillus sp. IW.에 의한 황화수소 제거

차진명,고한철,이인화 조선대학교 부설 유전생물공학연구소 1994 유전생물공학연구지 Vol.4 No.-

Thiobacillus sp. IW., capable of oxidizing hydrogen sulfide, was isolated from waste coal mine water around Hawsun in Chunnam province. The isolate was motile gram-negative rod shape, formed spore and grew up to be aerobically facultative chemolithotroph by using energy released from the oxidation of reduced inorganic sulfur compounds. It was found that the optimum thiosulfate concentration for growth was 32mM. The G+C content of the DNA was 65.0 mol%. The isolate had 16:1+17cyc, 16:0 as their major non-hydroxylated cellular fatty acids, 3-OH 12:0 as a hydroxylated fatty acid and also contained unidentified C_18 branched fatty acid. The ubiquinone system in the respiratory chain was Q-9. The cell productivity was 6.25mg/1.h and the isolate grew logarithmically up to 18 hour. The maximum specific growth rate of cell was 0.78h^-1 and generation time was 0.9 hour. In order to oxidize hydrogen sulfide in a bubble column reactor, Thiobacillus sp. IW. was immobilized on Polyvinylpyridine(PVP). 1.2l of broth medium containing 7.5g of immobilized PVP was used as a liquid phase and gas was supplied through the sintered glass distributor under hydrogen sulfide concentration controlled air stream. A Michaelis-Menten equation was applied for kinetics modeling and the maximum removal rate(V_m) and saturation constant(K_s) of hydrogen sulfide were 52.4(g-S/kg-dry PVP.d) and 62.88ppm, respectively.

형광성 Pseudomonas에서 Siderophore 생합성 유전자 탐색

김현희,고한철,여명구,김호상,서영우,박열 조선대학교 부설 유전생물공학연구소 1993 유전생물공학연구지 Vol.3 No.-

Kanamycin-sensitive Pseudomonads을 Tn5를 이용한 돌연변이 유도를 실시하여 kanamycin에 내성을 갖는 transconjugants를 선별하였고, siderophore 생합성하지 못하는 돌연변이체를 선별하기 위하여 MKB 평판배지에서 황색 hallow를 형성하지 못하는 colony를 선별하였다. 선발된 돌연변이체들의 genomic DNA애 Tn5가 삽입되었음을 확인하기 위하여 Southen blot hybridiz-ation을 실시하여 intact Tn5에 homology를 나타내는 밴드를 확인하였다. 각각의 변이주에서 밴드의 위치가 서로 다르게 관찰됨으로써 Tn5가siderophore 생합성 유전자에 무작위적으로 삽입되었음을 알 수 있었다. Gene library를 제조하기 위하여 wild typePY002의 genomic DNA를 Sau3A Ⅰ로 부분 절단하여 15-30kb 절편을 cosmid pLAFRS3에 접합하여 in vitro packaging 한 후 E. coli HB101얘 형질도입하여 P. fluorescens PY002의 genomic library를 재조하였으며, 이 library의 평균삽입 DNA 크기는 21kb였다. Pseudomonas fluorescens PY002, isolated from soil, was mutagenized with a transposon Tn5. The transposon Tn5 was randomly transposed into the chromosomal DNA of P. fluorescens PY002, as judged by Southem blot analysis with Dig-dUTP-labelled pSUP5011 (pBR325::Tn5) as probe. To construct a gene library from P. fluorescens PY002, the Sau3AI partial digested chromosomal DNA fragments of 15-30kb in sizes were ligated with BamHI-restricted cosmid vector pLAFR3. After the in vitro packaging, the resulting phages were transfected into E. coli HB101 and plated onto LBagar plate containing tetracyclin (12.5 ㎍/ml). The average sizes of insert DNA contained in the constructed gene library were approximately 21 kb in size range and the transduction number per 1 ㎍ of insert DNA was 9 x 10^3.

새싹 성장에 미치는 산소영향에 관한 연구

정경아,고한철 한국보건기초의학회 2017 한국보건기초의학회지 Vol.10 No.1

항산화란 활성산소(Oxygen free radical)의 산화 활동을 억제하거나 제거하는 것을 말한다. 산소는 두 가지 얼굴을 지니고 있다. 우선 산소는 우리 인간에게 없어서는 안 될 필수 불가결한 물질이다. 인간을 포함한 모든 동물은 몸 밖에서 영양분을 섭취하고 그것을 산소로 연계시켜 에너지를 얻는다(Zhang et al., 1992). 반면 우리 몸에는 끊임없이 활성산소(유해산소)가 생긴다. 활성산소는 이른바 불완전하게 연소된 산소. 전자를 잃어버려 항상 불안한 이 산소가 세포를 공격해 상처를 입히는데 이를 산화작용이라고 한다. 쇠가 녹슬 듯 세포가 병드는 것이다. 전문가들은 각종 질병의 90%와 노화 과정 등에 활성산소가 영향을 미치는 것으로 분석한다(Kim. et al., 1997; Talalay, et al., 1995). 인체는 생명을 유지하는 동안 끊임없이 반응성이 강한 산소화합물(reactive oxygen species; ROS)과 유리기를 생성함으로써, 산화물질(pro-oxidants)을 축적하게 된다.

녹농균의 항균제 내성양상 및 Rifampin과 Quinolone제와의 병합효과

박정평,고한철,나영천,임용,이현국,양남웅,박열 대한화학요법학회 1990 대한화학요법학회지 Vol.8 No.2

Clinically isolated 92 strains of Pseudomonas aeruginosa were serotyped and their resistance patterns to total 24 kinds of antimicrobial agents (i.e., Penicillins, aminoglycosides, cephalosporins, quinolones) were studied by MIC(minimal inhibitory concentration). In addition, serotype G(16 strains) and E(16 strains), which isolated with high frequency in clinical field, were used for a study about combined effects of quinolones and rifampin by the checkerboard method. These drugs are being spotlighted as therapeutics of Pseudomonas aeruginosa infections. The results were as follows; Serotypable strains of Pseudomonas aeruginosa were 87 strains(94.6%) out of 92 strains clinically isolated. In these strains, G types were isolated in largest numbers(28 strains, 30.4%), E types were 25 strains (27.2%), B types 12 strains (13.0%), the other types were less than 7% and J,K,N types were not typed at all. In the MIC peak of each antibiotics, ciprofloxacin showed lowest peak as 0.25㎍/㎖, another third generation quinolones also showed low MIC peaks of 8㎍/㎖ and less. Amikacin, the aminoglycosides, and rifampin revealed MIC as low as 32㎍/㎖ relatively, but the others did not. MIC of gentamicin, tobramycin, nalidixic acid, rifampin, piperacillin and amikacin were distributed in a wide range of 0.5㎍/㎖ to >512㎍/㎖, but in MIC_(50) of all above antibiotics were 256㎍/㎖ or more, and MIC_(90) were above 512㎍/㎖, the third generation quinolones except rosoxacin showed very low MIC ranges less than 64㎍/㎖, and revealed good antimicrobial activities as seen in that MIC_(50) were less than 2㎍/㎖ and MIC_(90) were below 8㎍/㎖. On seeing MIC ranges according to serotypes, in most antibiotics, E types were slightly resistant than G types but there are no much differences. In combinations of rifampin and quinolones, rifampin-enoxacin combination revealed relatively as good synergistic effect as 37.5%(12 strains), rifampin-nalidixic acid showed 34.4%(11 strains), and the other combinations also showed good results comparatively. In addition, rifampin-ciprofloxacin had antagonistic effect of 31.3%(10 strains). But on the whole, it can be said that these combinations have much affirmative significance in antibiotic therapy to Pseudomonas aeruginosa. According to the serotypes, synergism of rifampin-nalidixic acid to G types was zero%(0 strain) and 31.3%(5 strains) to E types. All combinations except rifampin-nalidixic acid revealed high synergistic effects to E types and in proportion to this, antagonistic effects were low. This high synergistic effects to E types were in a contrast to slightly high resistance patterns of E types.

Acinetobacter junii POH 의 페놀 생분해 특성

차진명,고한철 한국공업화학회 1998 응용화학 Vol.2 No.1

Acinetobacter junii POH was formed the yellow color colony on agar plate containing phenol as a vapor phase and grew well in a liquid minimal medium. the optimal concentration for phenol-biodegrading was 1000ppm. Acinetobacter junii POH was capable of growing on aromatic compound such as benzene, phenol, m-cresol, p-cresol, p-xylene and toluene but did not grow on o-cresol. With this strain, catachol was degraded through almost meta-cleavage pathway.

Acinetobacter junii POH 의 페놀 생분해 특성

차진명,고한철 한국공업화학회 1998 한국공업화학회 연구논문 초록집 Vol.1994 No.1

Acinetobacter junii POH was formed the yellow color colony on agar plate containing phenol as a vapor phase and grew well in a liquid minimal medium. the optimal concentration for phenol-biodegrading was 1000ppm. Acinetobacter junii POH was capable of growing on aromatic compound such as benzene, phenol, m-cresol, p-cresol, p-xylene and toluene but did not grow on o-cresol. With this strain, catachol was degraded through almost meta-cleavage pathway.