• KSBB Journal
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• 제5권3호
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• pp.269-277
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• 1990

메탄올이용균인 Methylobacillus sp. SKI의 균체생산효율을 증대시킬 목적으로 마이크로컴퓨터 제어 배양기를 이용해 고농도 유가배양을 수행하였다. 배양액의 초기 메탄올농도를 1.0%(v/v)로 하여 회분배양할 경우 배양 13시간만에 2.07g/l의 균체농도에 도달하였다. 공기 공급에 의한 유가배양에서 최대교반속도 1200rpm, 최대 공기유량 5.0$\ell$/min의 조건으로 배양시 약 15시간만에 13.7$\ell$/ldml 균체농도에 도달하였다. 산소공급에 의한 유가배양에서 최대교반속도 1200rpm, 공기유량 1.0$\ell$min, 최대산소유량 5.0$\ell$/min의 조건으로 배양시 17시간만에 45.3g/l의 균체농도에 도달하였다. 균의 대수 증식기를 공기공급에 의한 유가 배양으로 회분배양에 비해 약 3시간, 산소공급에 의한 유가배양으로 회분배야에비해 약 3시간, 산소공급에 의한 유가배양으로 회분배양에 비해 약 4시간 더 연장할 수 있었다. 즉, 유가배양로 단시간에 높은 농도의 균체를 얻은 것은 feedback제어에 의해 메탄올을 저해농도 이하로 유지시키면서 용존산소를 한계농도 이상으로 제어함으로써 균의 대수적 증식으로 연장시킨 결과이다. 산소공급에 의한 유가배양중 균체농도 27.6g/l에서 미량원소성분이 결핍되었고, 42.8g/l에서 $Mg^2^+$성분이 결핍되었으므로 배양중에 추가공급되었다.

• 한국미생물·생명공학회지
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• 제27권4호
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• pp.327-332
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• 1999

A highly productive fed-batch fermentation process was developed for the production of L-ornithine by using a new stabilized strain, Breviabcterium ketoglutamicum BK52. Fed-batch cultures with a continuous feeding of the complex medium were conducted on various operating conditions. The optimal concentration of phosphate in the complex medium was 2.1g/L. The optimal feeding rate of L-arginine was 0.028g/L/hr. The optimal feeding point of the complex medium was determined to be at 40 OD of the cell mass. The final L-ornithine concentrations within 64hrs of cultivation in 5 and 50 liter fermenters were 73g/L and 71g/L, respectively. The maximum overall L-ornithine productivity was 1.14g/L/hr which was about 2 times higher than that of the conventional fed-batch culture with intermittent feeding. The overall productivity of the fermentation system is remarkably improved by employing the optimized conditions, and it offers a significant potential for industrial application.

• Journal of Microbiology and Biotechnology
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• 제19권12호
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• pp.1603-1611
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• 2009

High-cell-density cultivation of yeast was investigated using the agricultural waste products corn steep liquor (CSL) and molasses. The Saccharomyces cerevisiae KV-25 cell mass was significantly dependent on the ratio between C and N sources. The concentrations of molasses and CSL in the culture medium were statistically optimized at 10.25% (v/v) and 16.87% (v/v), respectively, by response surface methodology (RSM). Batch culture in a 5-l stirred tank reactor using the optimized medium resulted in a cell mass production of 36.5 g/l. In the fed-batch culture, the feed phase was preceded by a batch phase using the optimized medium, and a very high dried-cell-mass yield of 187.63 g/l was successfully attained by feeding a mixture of 20% (v/v) molasses and 80% (v/v) CSL at a rate of 22 ml/h. In this system, the production of cell mass depended mainly on the agitation speed, the composition of the feed medium, and the glucose level in the medium, but only slightly on the aeration rate.

• Journal of Microbiology and Biotechnology
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• 제25권6호
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• pp.814-819
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• 2015

The natural crosslinking agent genipin has been applied widely in biomedicines and foods nowadays. Because of the special hemiacetal ring structure in its molecule, it can only be prepared by hydrolysis of geniposide according to biocatalysis. In this research, strategies including aqueous-organic biphasic catalysis and substrate fed-batch mode were adopted to improve the biocatalysis process of genipin. A 10 L ethyl acetate-aqueous biphasic system with geniposide fed-batch led to a satisfying genipin yield. With Fusarium solani ACCC 36223, 15.7 g/l genipin in the ethyl acetate phase was obtained, corresponding to space-time yields of 0.654 g l^-1 h^-1.

• Journal of Microbiology and Biotechnology
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• 제10권3호
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• pp.321-326
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• 2000

To examine the feasibility of the long-term production of the human granulocyte colony stimulating factor (hG-CSF) using the $_{L}-arabinose$ promoter system of Escherichia coli, flask relay culture and cyclic fed-batch culture were performed. In the flask relay culture, it was found that the pismid was maintained stably up to about 170 generations in an uninduced condition, whereby the cells could also maintain the capability of expressing hG-CSF expression were maintained stably up to at least 100 generations. In contrast, in the cyclid fed-batch culture, segregational plasmid instability was observed within about 4 generations after induction, even though the cell growth and hG-CSF production reached their maximum balues, 78.0 g/l of dry cell weight and 7.0 g/l of hG-CSF, respectively. It would appear that, when compared to the flask relay culture, the high-cell density and high-level expression of hG-CSF in the cyclic fed-batch cultrure led to the segregational plasmid instability; in other words, a severe metabolic burden existe on the cells due to the high-level expression of hG-CSF. Accordingly, based on these long-term cultures, the segregational and structural plasmid instability was observed and a strategy to overcome such problems could be designed.

• KSBB Journal
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• 제29권4호
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• pp.303-306
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• 2014

Antifreeze proteins (AFP) inhibit ice growth to permit the survival of polar organisms in the cold environments. The recombinant AFP from an Antarctic bacterium, Flavobacterium frigoris PS1, FfIBP (Flavobacterium frigoris ice-binding protein), was produced using Pichia pastoris expression system. The optimum fermentation temperature ($30^{\circ}C$) and pH (5) for FfIBP production were determined using a fed-batch culture system. The maximal cell density and purified FfIBP were 112 g/L and 70 mg/L, respectively. The thermal hysteresis (TH) activity (0.85) of FfIBP obtained using a glycerol-methanol fed-batch culture system was 2-fold higher than that of the LeIBP (Leucosporidium ice-binding protein). This work allows for large-scale production of FfIBP, which could be extended to further application studies using recombinant AFPs.

• Journal of Microbiology and Biotechnology
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• 제7권5호
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• pp.323-328
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• 1997

The recombinant Escherichia coli BL21(DE3)pLysE : pTCGT1 was grown to overproduce Bacillus macerans cyclodextrin glucanotransferase (CGTase) able to synthesize ${\alpha}$-cyclodextrin (CD) with a selectivity of 67%. A number of batch fermentations were performed to test the possibility of using lactose as an inducer of the E. coli T7 promoter system. A mixture of isopropyl ${\beta}$-D-thiogalactoside (IPTG) and lactose (1 : 1) gave a maximum CGTase activity of 2.4 U/ml, which was higher than the value obtained with induction by IPTG alone. Fed-batch fermentations involving a glucose-controlled growth period followed by a gene-expression phase with mixtures of IPTG and lactose were employed to achieve high cell density and thereby increase total CGTase activity. Optimized fed-batch fermentation using the modified inducer (IPTG : lactose=1 : 3) and 100 g/l yeast extract solution in the gene-expression phase resulted in a maximum CGTase activity of 62.9 U/ml and a final cell mass of 53.5 g/l, corresponding to a 31-fold increase in CGTase activity and a 29-fold increase in cell mass compared with the control batch fermentation.

• KSBB Journal
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• 제15권2호
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• pp.174-180
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• 2000

본 연구에서는 다른 host cell 에 비하여 여러 가지 장점을 가지고 있는 Methylotrophic yeast 중 Pichia pastoris와 Hans-enula polymorpha의 fed batch 실험을 통하여 유전자 재조합 단백질 발현최적조건을 구하여 각 균주의 유전자 재조합 albumin 발현 최적화 연구를 수행하였다. 글리세롤이 두 균주의 promoter의 AOX 1과 MOX promoter repression에 미치는 영향을 확인한 바 H. polymorpha가 P. pastoris보다 promoter repression이 심함을 알 수 있었다. 두 균주의 promoter를 induction시키는 최적 메탄올 농도는 P. pastoris의 경우 메탄올 8g/L, H. polymorpha의 경우는 13 g/L임을 알 수 있었다. 또한 메탄올에 의한 induction시기는 두 균주 모두 O.D. 4 정도되는 exponential growth stage에서 메탄올을 첨가하는 경우가 초기 세포 성장단계에 메탄올을 첨가한 경우에 비해 약 20% 정도 높아짐을 확인하였다. 두 균주의 재조합 albumin 발현에 미치는 pH의 영향을 조사하였는데, p. pastoris의 경우 pH 5에서 가장 높은 albumin 생산성을 보여 약 300mg/L albumin을 발현하였고, H. polymorpha 의 경우 pH 5와 6에서 최대 약 180 mg/L의 albumin을 발현하였지만 pH 8에서는 이의 절반 수준에 그쳤다. 두 균주의 최적 fed-batch 방법을 확인하는 실험을 수행하였는데 P. pastoris의 경우의 최적 fed-batch 방법은 mixed feeding은 바람직하지 않고 글리세롤 배지를 공급하여 세포를 성장시킨 후 글리세롤 공급을 멈추고 바로 메탄올로 전환하는 방법을 효과적이며, H. polymorpha의 경우 비성장속도 제어를 통한 글리세롤 공급으로부터 메탄올 공급으로의 단계적 전환방법이 균주의 albumin 발현에 큰 영향을 준다는 것을 알 수 있었다. 이 방법을 통하여 두 균주의 고농도 배양 실험을 수행한 결과 P. pastoris의 경우는 O.D. 300에서 약 4.7g albumin/L를 발현하였다. 이와같은 결과를 바탕으로 산업체에서 methlotrophic yeast를 이용한 상업화를 계획함에 있어서 host로서의 균주를 선택할 수 있는 기본 자료를 제공함과 아울러 균주가 선택된 후에 그 균주를 이용한 재조합 단백질 최적화 방법을 제공하여 줄 것으로 생각된다.

• Journal of applied mathematics & informatics
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• 제24권1_2호
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• pp.387-397
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• 2007

According to the controllability of pulse times and the amount of jumps in the states at these times in the process of fed-batch culture producing 1,3-propanediol, this paper proposes a terminal optimal control model, whose constraint condition is the nonlinear impulsive delay system. The existence of optimal control is discussed and an optimization algorithm which is applied to each subinternal over one cycle for this optimal control problem is constructed. Finally, the numerical simulations show that the terminal intensity of producing 1,3-propanediol has been increased obviously.

• 한국미생물·생명공학회지
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• 제20권6호
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• pp.668-676
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• 1992

Alcaligenes eutrophus에 의한 poly-Beta-hydroxybuty-rate(PHB) 생산을 최적화하기 위해서 1단계에서는 성장을 최대화하고, 2단계에서는 생산을 최대화하는 이단 유가식 배양을 채택하였다. 성장단계에서 탄소원과 질소원의 최적농도는 회분식 배양 모델로부터 결정되었는데, 각기 16.64g/l와 0.54g/l이었다. 생산단계에서 질소원의 경우, 유가식 배양실험을 통해 0.07g/l가 최적인 것으로 나타났다. 탄소원의 최적농도는 모사로부터 결정되었고 이때 최적 전환시간도 함께 결정되었다. 생산공정의 제어를 위하여 시간지연을 고려한 적응비례 되먹임제어를 이용한 결과, 탄소원 농도 및 질소원농도를 최적의 상태로 유지시킬 수 있었다.